W heal x Rye = T rilicale

"that 11 may g1L'<' seed IQ thl! ~0 11-er . and bread 10 the cater" [/sai(lltj IDRC-02le

Nutritive Value of Triticale Protein (and the proteins ol wheat and rye)

Joseph H. Hulse Director. Agriculture. Food and Nutrition Sciences International Development Research Centre

Evangeline M. Laing Little Chalfont, Bucks, England

UDC 612.398:633.I © 1974 International Development Research Centre Head Office: Box 8500, Ottawa, Canada. KIG 3H9 Microfiche Edition $1 Abstract Triticale promises higher yields of grain than the most productive bread wheats. Advanced triticale lines are at least equal in protein content and superior in biological quality to wheat. Triticale has a higher content of the essential amino acid, lysine, which is first limiting for triticale, wheat, and rye. It is expected that further im­ provements in biological value will follow, particularly from a more intensive search for genetically superior varieties of rye. In addition to increasing protein quality by genetic control, lysine content of triticale foods can be improved by supplementation with other materials com­ paratively rich in lysine, the food legumes offering the most attractive source for the poorer nations. The considerable literature devoted to the processing and supple­ mentation of wheat and wheat flour is largely relevant to triticale. While more reliable data based upon advanced triticale lines are needed, present evidence suggests that sound, healthy triticale is equal, and possibly superior, to wheat in most farm animal diets. Since triticale will tolerate environments un­ suitable for wheat, it promises a significant addition to the world's cereal crops for both human food and animal feed. Triticale 's ultimate contribution cannot be fully realized without a closer collaboration than is customary among plant scientists, nutritionists, and food scientists and technologists. In particular, there is an urgent need for well-defined and universally accepted standards and methods of chemical analysis and biological evaluation by which to select those triticale lines which are genetically superior in protein quality.

Resume Les rendements en grain du triticale semblent devoir etre plus eleves que ceux des cereales panifiables Jes plus productives. Les lignees de multiplication du triticale ont une teneur en proteines au moins egale a celle du ble et Jui sont superieures sur le plan des qualites biologiques. La teneur du triticale en lysine, l'acide amine essentiel qui constitue le premier des facteurs limitants du triticale, du ble et du seigle, est plus importante que chez Jes autres cereales. L'avenir devrait amener une nouvelle amelioration de sa valeur sur le plan biologique, en particulier a la suite de recherches plus poussees sur des varietes de seigle genetiquement superieures. En plus d'une amelioration des qualites proteiques du triticale grace a la maitrise de sa genetique, ii est possible d'ameliorer la teneur en lysine des aliments qui en sont tires en Jes complementant avec d'autres matieres premieres comparativement riches en cet element: Jes legumineuses alimentaires constituent dans ce domaine la source la plus interessante pour Jes pays defavorises. Une tres grande partie de la masse de textes consacres a la transformation et a la complementation du ble et de la farine de ble est applicable au triticale. Bien que l'on ait besoin de donnees plus sfires fondees sur l'etude des lignees de pointe du triticale, Jes donnees dont on dispose deja laissent a penser que pour la plupart des regimes alimentaires des animaux d'elevage, un triticale de bonne qualite est un composant au moins egal sinon superieur au ble. Etant donne que le triticale tolere des conditions de milieu que ne supporterait pas le ble, ii constitue un element riche de promesses pour un accroissement des disponibilites mondiales en cereales destinees a l'alimentation des hommes et des animaux. II ne sera pas possible d'etablir pleinement la contribution finale que peut apporter le triticale, sans un renforcement de la collaboration dans ce domaine entre Jes phytotechniciens, Jes nutritionnistes et Jes specialistes et Jes techniciens de l'industrie alimentaire. II est necessaire notamment de definir rapidement et avec precision des normes et des methodes universellement acceptees d'analyse chimique et d'evaluation biologique permettant de selectionner des lignees de triticale genetiquement superieures sur le plan des qualites proteiques.

2 Contents

AUTHORS' PREFACE 5

CHAPTER 1. BACKGROUND AND SUMMARY 7 Triticale: Its Nature and History 7 The CIMMYT-Manitoba Project 7 Cereal Grains in Human Nutrition 8 Nature and Composition of Cereal Protein 10 Scope of the Text and Summary 12 Evaluation of Plant Proteins 12 Varietal and Environmental Factors 16 Nutritional Inhibitors and Toxic Factors 19 Processing and Supplementation 20

CHAPTER 2. METHODOLOGY FOR EVALUATION Of PLANT PROTEINS FOR HUMAN USE 27 Need for Methods 27 Total Protein Analysis 28 Amino Acid Analysis 29 Protein Hydrolysis 29 Ion Exchange Chromatography 29 Microbiological Assays 29 Amino Acid Units 29 Available Amino Acids 29 Chemical Score 30 Amino Acid Availability 31 DNFB Method 31 Enzymatic Hydrolysis 32 Microbiological Methods 32 Effect of Heat Processing 32 Biological Quality 32 Protein Efficiency Ratio 32 Net Protein Retention 33 Net Protein Utilization 33 Slope Ratio Assay 33 Bioassays-Conclusions 34 Rapid Screening Methods 34 Microbiological Methods 34 Dye-Binding Methods 35 Biological Methods 36 Rapid Screening-Conclusions 36 Bioassays with Humans 36 Suggested Testing Procedures 37 Summary 37 Recommended Methods 38 PER Assay 38 NPR Assay 38

3 Contents (concluded)

CHAPTER 3. GENETIC, VARIETAL, ENVIRONMENTAL AND AGRONOMIC FACTORS 41 Introduction 41 Genetic and Varietal Factors 41 Environmental and Agronomic Factors 44 Triticale 45 Genetic and Varietal Effects 45 Environmental and Agronomic Effects 69 Wheat 70 Genetic Effects 70 Varietal Influence 72 Environmental Influences 77 Agronomic Influences 81 Wheat and Rye Compared 88 Genetic and Varietal Effects 88 Environmental and Agronomic Influences 89 Rye 90 Genetic and Varietal Effects 90 Environmental and Agronomic Influences 92

CHAPTER 4. NUTRITIONAL INHIBITORS AND TOXIC FACTORS 95 Introduction 95 Naturally-Occurring Inhibitors 95 Resorcinols 95 Phytic Acid 97 Enzyme Inhibitors 99 Growth-Depressing Factors 100 Toxic Substances IOI Ergot IOI Aflatoxin I 02 Tolerance Levels 103 Processing Additives I 03

CHAPTER 5. PROCESSING AND SUPPLEMENTATION WITH OTHER PROTEIN SOURCES 105 Introduction I 05 Milling 105 Secondary Processing I 09 Supplementation with Other Protein Sources 109 Milling 110 Triticale 110 Wheat Flour 113 Rye Flour 122 Secondary Processing 124 Triticale 124 Wheat and Rye 127 Supplementation with Other Protein Sources 134 Synthetic Amino Acids 134 Cereal Proteins 143 Egg and Milk Protein 145 Food Legume and Oilseed Protein 147 Protein from Microorganisms 151 Fish Proteins 152 Conclusion 156

EPILOGUE 158

REFERENCES 159

INDEX 181

4 Authors' Preface

Cereal grains are the main source of calories and protein for many people in the developing world. Triticale is a man-made cereal grain, a cross between wheat (Triticum spp.) and rye (Secale spp.). This publication com­ prises a review of the literature, a commentary, and a bibliography, relative to the biological value of triticale, and its parents wheat and rye. It concerns itself predominantly with the proteins of these cereals. It is hoped that it will prove interesting and useful to plant scientists, nutritionists, food scientists, and technologists, and others, particularly in developing countries, where triticale may be grown and eaten. The present state of triticale development is largely the outcome of the Triticale Project, a scientific cooperative venture between the International Maize and Wheat Improvement Centre (CIMMYT) and the University of Manitoba, supported by the Canadian International Development Agency and the International Development Research Centre. The authors wish to acknowledge, with admiration, the work of many plant scientists, particularly those at CIMMYT and the University of Manitoba, whose imaginative curiosity and scientific skill have made possible the advanced state of develop­ ment of this remarkable new cereal grain. In the preparation of this manuscript particular thanks are due to Ors J. M. Mclaughlan and J. A. Campbell who kindly contributed Chapter 2, Methodology For Evaluation of Plant Proteins For Human Use. The help and facilities extended by Dr C. T. Greenwood, Director of the Flour Milling and Baking Research Association, Dr P. C. Spensley and his colleagues at the Tropical Products Institute, and Prof. J. B. M. Coppock are also acknowl­ edged with appreciation. The helpful editorial contribution of Mr R. L. Macintyre, and the assistance of Miss Wanda Huff, Mrs Margaret Kovesi, Miss Caroll Rathwell, and Mrs Helen Gay, who typed the manuscript are gratefully acknowledged.

November 1973 JosEPH H. HuLsE EVANGELINE M. LAING

5 Acres of lriticole growing at CIM M YT. El Bown. Mexico Chapter 1

BACKGROUND AND SUMMARY

Triticale: Its Nature and History food and nutrition scientists, and cereal tech­ nologists interested in triticale, and in particular Dr. Norman E. Borlaug, Nobel Peace Prize to both scientists and agricultural policy-makers Winner in 1970 and head of the international in developing countries where triticale might wheat research program at the International Maize beneficially join the existing spectrum of estab­ and Wheat Improvement Center (CIMMYT), lished cereal grains. Mexico stated in a paper published in October Triticale is not as new as is sometimes suggested. 1973 " ... it appears certain that triticales will soon It has existed as an academic curiosity since the add to the world's food production potential. first naturally occurring triticale was described in Because of their good protein and amino acid a report to the Botanical Society of Edinburgh in properties, they may also play a role in correcting 1875. When a natural cross is made between wheat protein malnutrition amongcereal-eatingnations." and rye the first generation (Fi) plant is normally Triticale is an artificial genus synthesized by sterile. It was discovered in 1937 however that combining the genomes of wheat (genus Triticum) when an F seedling was treated with colchicine and rye (genus Secale). If the wheat parent is a 1 its chromosome makeup was doubled and the tetraploid (T. durum), the resultant triticale will resultant triticale plant was partially fertile. This be hexaploid (Triticale hexaploide); if the wheat is discovery opened the door to the genetic improve­ hexaploid (T. aestivum), the resultant triticale will ment which has since taken place. Colchicine is be octaploid (Triticale octaploide), the rye parent an alkaloid present in certain lilaceous plants of in each case being diploid. So far the hexaploid the genus Colchicum which includes the autumn triticales have proved more stable than the crocus. octaploids. It should be emphasized that triticale Since triticale is a cross between wheat and rye, is not a single species. Triticale is a genus and it would be expected to inherit characteristics like wheat and rye, it embraces many cultivars of from both of its parents. While this is so, triticale widely variable characteristics. is, without question, a unique plant, and the main The early history of triticale research and purpose of this publication is to review what is development was described by Zillinsky and known of the factors which influence the biological Borlaug (197la), and Zillinsky (1973) has pre­ value of its protein. Since triticale may derive sented a more recent review. A description of varying degrees of genetic influence from its progress from many parts of the world, based parents, it was considered necessary to review as upon the proceedings of a workshop sponsored much of the literature relating to the biological by the International Development Research Centre value of wheat and rye as appeared relevant and held at CIMMYT in October 1973 will be pro­ potentially useful to the future nutritional im­ duced as an IDRC publication. provement of triticale. This publication seeks to bring together what is known about the biological value of the protein of triticale, and as much as appears relevant con­ The CIMMYT -Manitoba Project cerning the biological quality of the proteins of Triticale research began in Canada at the Uni­ its parents wheat and rye. It is hoped that the versity of Manitoba where in 1954 the Rosner publication will be helpful to all plant scientists, research chair was endowed by the Samuel and 7 8 MONOGRAPH IDRC-02le

Saidye Bronfman Family Foundation. In 1963 the cytogenetics, biochemistry, plant physiology and Manitoba triticale program was extended into a pathology, and cereal technology. nursery at the CIANO Research Station at Ciudad It is doubtful if any group of plant scientists is Obregon, Mexico. In 1965, using the University better aware than the CIMMYT team of the of Manitoba's material as initial breeding stock, potential for increased productivity in cereal a substantial triticale program was initiated by grains. The Food and Agriculture Organization the Centro Internacional de Mejoramiento de (I 970a) data indicate that the yields of wheat in Maiz y Trigo (CIMMYT) in Mexico, which sub­ Mexico increased from 8.8 to 28.4 kg per hectare sequently has grown into a fruitful cooperative between 1948-52 and 1970. Dalrymple (I 972) research program between CIMMYT and the estimates that the area under high yielding University of Manitoba. varieties outside Mexico rose from 9,000 to more This cooperative effort has flourished for than IO million between 1965/66 and 1970/71. several reasons. First, the triticale program bene­ fited from CIMMYT's highly organized, intensive Cereal Grains in Human Nutrition and successful bread and durum wheat research. Wheat is grown over a wide area of the world's The genetic base of triticale was diversified and its arable land but is generally most successful qualities improved both through a planned pro­ between the latitudes 30° and 60° North and 27° gram of crossing to CIMMYT's immense pool of and 40° South. It has been known to grow at wheat varieties and also by an adventitious, altitudes from sea level up to 15,000 feet and at promiscuous outcrossing which gave rise to the precipitations between 275 and 760 mm. It grows fertile triticale progeny known as Armadillo. best in temperate and subtropical climates and Second, two generations of triticale could be prefers heavy loam and clay soils (Kent 1966). grown each year, one in Mexico, one in Canada. Close to 220 million hectares of the earth's sur­ Third, alternate generations could be grown under face are given over to the growing of wheat, the widely different environmental conditions, thus total world production being of the order of 343 enabling the identification of lines with day-length million metric tons per annum. Roughly 20 insensitivity and wide adaptability. million hectares are under rye for a total world The close working relation between CIMMYT production of about 31 million metric tons. No and the University of Manitoba gave rise to the official figures for the commercial production Triticale Project now supported by the Inter­ of triticale are readily available but it is estimated national Development Research Centre and the that about 500,000 hectares were planted for Canadian International Development Agency. harvesting in 1971 (Zillinsky and Borlaug 1971 b). The objective of the Triticale Project is to develop Triticale testing nurseries are to be found in 52 triticale varieties that will complement wheat, different countries. barley, rye and other cereal grains and to provide a The number of known wheat varieties is very new crop well adapted to a wide variety of con­ large indeed, more than 16,000 in the World ditions, a crop capable of substantially increasing Collection having been screened for protein and food production in the developing countries. lysine content (Johnson and Mattern 1972). The Because the primary objective of the Triticale principal wheats grown are the hexaploid Triticum Project is to improve the nutritional welfare of aestivum used mostly in bread and baked products, the developing countries, serious attention is and the tetraploid Triticum durum which, in the being given to the grain's biological value. western world, is particularly popular for ali­ While both CIMMYT and the University of mentary pastes such as noodles and macaroni. Manitoba are cooperating in plant breeding The most agronomically successful varieties of research, the primary responsibility for the de­ triticale so far developed are the hexaploid velopment of improved breeding lines of triticale derivatives of Triticum durum crossed with the rests with CIMMYT, where, under the direction diploid rye Secale cerea/e. of Dr. Norman E. Borlaug, the high yielding Considerably fewer varieties of rye than wheat dwarf varieties of wheat were developed and dis­ are documented. Rye will grow in roughlY the seminated to many parts of the world. The same latitudes as wheat but is much more tolerant scientists at the University of Manitoba are of sandy soils and soils of lower fertility (Shands addressing themselves to fundamental studies in 1969). Consequently, it is hoped the triticale HULSE/LAING: PROTEINS OF TRITICALE 9

TABLE I. Proportion of daily per ca put nutritional supply from different sources, 1962 and proposed 1985 levels (data from Food and Agricultural Organization 1969a).

Africa S. of Sahara Asia Latin America Near East

Calories• Proteins• Calories Proteins Calories Proteins Calories Proteins

1962

Cereals 51.2 53.2 65.9 59.9 40. 7 40.0 65.9 67.0 Pulses", nuts, seeds 8.9 18.3 9.4 20.0 6.7 15.7 3.7 7. I Vegetable oils 4.6 4.1 5.2 5.2 Other food crops 29.7 12.8 14.3 6.7 30.5 8.0 15.0 3.1 Total plant products 94.4 84.3 93.7 86.6 83. I 63.7 89.8 79.7

1985

Cereals 51. 3 51.4 61.0 57.5 39.7 39.4 62.6 64.5 Pulses, nuts, seeds 9.4 19.3 9.7 21.4 6.7 16. I 4.0 8.0 Vegetable oils 4.9 6.0 6.5 5.9 Other food crops 28.4 12.1 17.3 7.3 30.2 8.1 17.0 6.0 Total plant products 93.9 82.2 94.0 86.2 83. I 63.6 89.5 78.5

•All data are estimated average percentages of total diet. hPulses are dried legumes. It is possible that in this table the term "pulse" includes food legumes in general. hybrid will demonstrate a greater environmental productive unions, close cooperation between the adaptability than either of its parents and, in two having been a rare phenomenon. Within the particular, that it will grow successfully on what Triticale Project, however, a close working partner­ may be described as marginal lands. ship does exist between the plant scientist and the As will be detailed later in the text (Table 3), nutritional biochemist. It is hoped therefore that, the protein of rye is generally higher in the in addition to its technical output, the Triticale essential amino acid lysine than is wheat and Project will serve as a model for other future plant therefore triticale varieties superior in nutritional improvement projects. value to the common wheat varieties can be syn­ The Triticale Project is designed primarily for thesized. Wheat, on the other hand, is much more the benefit of people in the less developed countries widely consumed than rye in human diets. In of the world, particularly those who are below, or Asia, for example, based upon data given in the close to, the margin of dietary calorie and protein FAO Production Yearbook 1970, per capita sufficiency. One cannot overstate the importance production of wheat increased from 26 kg to 38 of cereal grains and grain legumes in human diets, kg. The world production and consumption of particularly in those of the world's poor and mal­ rye is of a much lower order of magnitude. nourished. Cereals, generally, make a larger The Triticale Project seeks to offer a cereal single contribution to both energy (calories) and grain technologically and gustatorily as attractive protein in the human diet than any other single as wheat, with a biologically superior protein commodity group. Consequently any significant derived from its rye ancestry. improvement in the biological value of cereal In "Food and the New Agricultural Tech­ protein will add significantly to the quality of nology", Palmer (1972) writes about "The final the diet. Table I, taken from the FAO Provisional divorce of agriculture and nutrition through the Indicative World Plan for Agricultural Develop­ agency of the 'Green Revolution'." Until recently, ment (Food and Agriculture Organization I 969a), the working relations between plant scientists and states the approximate proportion of calories and nutrition scientists might better be described as protein supplied by cereals, food legumes and casual transitory affairs rather than durable other crops to the average diets of the peoples of 10 MONOGRAPH IDRC-02le

Africa, Asia, Latin America and the Near East. from 1.2 to 0.9 kg and lentils from 0.6 to 0.4 kg. The data include estimates for the year 1962 and Dalrymple and Jones (1973) state that .. The drop the forecast for 1985. in pulses [production in India] reflects a long term The predominance of cereal grains and grain decline in production which preceded the ·green legumes as sources of both calories and protein revolution' and which is a function of low yields is readily evident. It is equally evident that cereal and low prices." and food legumes are expected to provide most If these trends persist, one can anticipate that of the calories and protein through the year 1985 the poor people of the developing countries may and beyond. The authors of the Indicative World have to depend upon cereals for an even higher Plan forecast that between 1965 and 1985 the proportion of their future protein calories. Con­ population of the poor countries is expected to sequently, concern must be given both to the increase by about 60%. The population pressure, qualitative and the quantitative aspects of cereal added to other demand pressures, suggest to the protein production in the less developed countries. authors of the Indicative World Plan that the While it is not suggested that a nutritionally production of cereal grains and other crops needs superior triticale plant will solve the protein to increase by no less than four percent per annum. problem of all or even most of the world's mal­ At its 59th meeting in December 1972, the Council nourished, triticale can, and hopefully will, pro­ of FAO expressed "grave concern" over the trend vide an additional valuable source of good quality in agricultural production which showed a global protein and calories. increase of only one to two percent in 1971 against Any protein source must be considered in a target increase of four percent. relation to the total diet of the specific population The advent and introduction of the high-yield­ for which it is intended. Conclusions based upon ing varieties of wheat and rice gave cause for average data for large populations should be cautious optimism that, given reasonably favour­ viewed with caution since significant variations in able climatic conditions, the total production of both the quantity and quality of the protein in cereal grains in the developing countries could be the diet are known to occur among different significantly increased. However, the tragic events groups within any given population, among which occurred in 1973 in West Africa and other families within a group, and even among members parts of the world serve to emphasize how tenuous within a family. is the present balance between human need and Differing solutions will be needed for different the availability of the major subsistence crops. population groups and, therefore, this review In addition to this precarious calorie balance, addresses itself not only to the biological improve­ there is little evidence to suggest that the overall ment of the triticale plant but also to the beneficial nutritional quality of the diet, and in particular and detrimental influences of processing and of its protein content, is improving to any marked supplementation. The Triticale Project is in fact degree among the developing nations. The data as much concerned with how triticale will be used quoted in Table 1 are overall averages covering as with how it will be grown. While little is large regional populations; it is probable that the recorded about the processing of triticale, the poorest and the most malnourished people of literature on wheat and rye processing and sup­ these regions derive a significantly higher pro­ plementation is extensive and relevant. It is portion of their calories and protein from cereal hoped, therefore, that this aspect of the review grains and grain legumes than the· means quoted. will be helpful to the improved future use of Cereal protein and legume protein, when eaten triticale. together, provide a more nutritious diet than either eaten separately. There is evidence to suggest Nature and Composition of Cereal Protein that because of the higher cash return per hectare The name "protein" is etymologically derived from cereals, the areas allocated to grain legumes, from the Greek npwr:e1ou meaning "primary" particularly in Southeast Asia, are contracting. and is so named because protein is the primary Palmer (1972) quoting data cal.culated from FAO and fundamental material of most living organ­ Production Year Book 1970, indicates that in isms. "Protein" is the name given to a class of Asia between 1948-52 and 1970 per capita chick­ organic compounds made up of linked amino pea production declined from 6 to 3 kg, soybeans acids composed of nitrogen and other elements. HULSE/LAING: PROTEINS OF TRITICALE II

Amino acids are the essential building materials while the germ and scutellum fractions contain of which all animal tissues and organs are com­ more protein than any other fraction, they repre­ posed and maintained. sent so small a proportion (2.5%) of the whole Protein is essential to the growth and restoration grain that their contribution to the total protein of body tissue and is particularly important in the content is indeed small. On the other hand the diets of young children, pregnant and lactating endosperm plus the surrounding aleurone layer women, and to aid recovery after serious illness or provide approximately 88% of the total protein injury. There is evidence to suggest that serious present in the whole grain. Consequently any protein deficiency in early childhood can impede research aimed at either increasing total protein brain development and learning ability (Winick or improving the biological quality of the protein and Rosso 1969, 1970). should concentrate upon the storage proteins in Protein is an essential component of a balanced the endosperm and aleurone. Milling techniques, diet which must also be adequate in calorie sources. in particular, should strive to include as much of In a diet deficient in energy sources, the protein the aleurone layer as possible. A labelled diagram present may be used as a source of calories thus of a sectioned grain appears in Chapter 5 (Fig. 2). leading to an aggravated state of protein mal­ The early lines of triticale, in common with the nutrition (Carpenter 1970). The point of progeny of other wide genetic crosses, demon­ Carpenter's comment is that where both dietary strated a high frequency of shrivelled grains in calorie and protein deficiencies occur, the protein which the embryo and bran represented a rela­ will be consumed as an energy source thus ag­ tively high proportion of the grain. Since variability gravating the organism's protein deficiency as in the ratios of endosperm to bran and germ may, indicated by body nitrogen balance. It should be in themselves, make for variability in grain noted however that while an inadequate intake of nitrogen content and biological value, it is im­ protein can be seriously detrimental to health, no portant in reporting protein and amino acid harmful effects have been demonstrated from compositions in whole cereal grains to describe protein intakes in excess of probable need apart, the overall physical characteristics of the grain of course, from the hazards associated with under discussion. excessive overeating. Different proteins are composed of different The protein content of wheat has been shown combinations of amino acids. Some of these to vary widely among different varieties grown amino acids can be synthesized by living organisms under different environmental conditions. As from other nitrogenous material, the amino acids early as 1889 it was reported that protein contents which can be synthesized varying among different ranged from 7% in wheat grown in Scotland to higher animals. Other amino acids essential to the more than 24% in wheats grown in the Caucasus. diet cannot be synthesized in vivo but must be Though the precise accuracy of the data is doubtful ingested as such. These are known as the "essential" in the light of modem analytical methods, the or "indispensible" amino acids and must all be wide variability in total wheat nitrogen content is provided by the food eaten. Where all essential supported in more recent publications. Johnson amino acids appear together in appropriate and Mattern ( 1972) in their analysis of samples proportions in a single protein source, that source of bread wheat from the World Collection found is sometimes described as an "ideal" or "perfect" protein contents to vary from 6.0% to 23.0% with source of protein. As is described in greater detail a mean of 12.9% on a dry weight basis. The pro­ below, where a particular protein is quantitatively tein content of rye is believed to vary from about deficient in one or more amino acids, its nutritional 6.5% to close to 14.5%, the higher values originat­ or biological value can be improved by the ing mainly in North America (Kent-Jones and addition of the amino acid in which it is most Amos 1967). deficient. As is discussed below, both the total nitrogen Of the eighteen amino acids most commonly content and protein composition vary significantly found in natural edible protein, according to among the endosperm, seed coats (bran) and Kasarda et al. (1971), ten are generally described embryo (germ) of cereal grains. From the data in as essential for human infants though only nine Table 72 in Chapter 5, calculated from data pub­ may be essential for adults. Chickens and turkeys lished by Hinton (1953) for wheat it is evident that need a total of twelve amino acids. Rats are 12 MONOGRAPH IDRC-021e

TABLE 2. Pattern of amino acid requirements as milligrams per gram protein compared with milk protein (data from World Health Organization 1973).

Suggested patterns of requirement Reported composition

Schoolchild Human Cow's Amino acid Infant 10-12 Years Adult Milk Milk

Histidine 14 26 27 Isoleucine 35 37 18 46 47 Leucine 80 56 25 93 95 Lysine 52 75 22 66 78 Methionine+ Cystine 29 34 24 42 33 Phenylalanine + Tyrosine 63 34 25 72 102 Threonine 44 44 13 43 44 Tryptophan 8.5 4.6 6.5 17 14 Valine 47 41 18 55 64 Total 373 326 152 460 504 believed to require the same ten essential amino data being derived from an FAO publication acids as human beings and are therefore generally based upon a review of a large number of analyses used as test animals in determining the biological (Food and Agriculture Organization 1970b). The efficiency of protein sources intended for humans. table also cites what the FAO compilers have Altschul (1965) has reviewed the amino acid­ stated to be the first and second limiting amino compositions of proteins as they relate to biological acids for wheat, wheat products and rye. In values, and the FAO/WHO Ad Hoc Expert common with wheat and rye the first limiting Committee on Protein and Energy Requirements amino acid in triticale is lysine (Knipfel 1969; (World Health Organization 1973) has suggested Kies and Fox 1970b). Consequently where plant patterns of ten essential amino acid requirements breeding, processing and supplementation re­ for school children and adults. It is recommended search have been related to an improved biological by the Expert Committee that the additional quality, greatest attention has been given to amino acid, histidine, is required in infant diets. increasing the proportion of lysine. The FAO/WHO recommended levels appear in Table 2 wherein the higher concentration of essential amino acids per gram of nitrogen re­ Scope of the Text and Summary quired in the diets of infants and children is immediately evident. Following this introductory chapter, the text The biological value of a protein depends upon consists of a review of the relevant literature its content of essential amino acids in relation to under four chapter headings. The remainder of the requirement for these same essential amino this chapter comprises a summary of and com­ acids in the species of animal for which the protein mentary upon each of the four succeeding is intended. The essential amino acid in which a chapters. protein is most seriously deficient is known as the first limiting amino acid. Evaluation of Plant Proteins In addition to the essential amino acids, humans Chapter 2 has been kindly contributed by Dr. and other animals require non-specific sources of J. M. Mclaughlan and Dr. J. A. Campbell of nitrogen from which to synthesize the "non­ Health and Welfare, Canada. Dr. Campbell was essential" amino acids needed for body building the Chairman and Dr. Mclaughlan a member of and maintenance. IDRC's Working Group on Biological Evalootion Table 3 quotes the average amino acid contents and both are internationally recognized authorities of wheat, wheat products, rye and triticale, this on the nutritional evaluation of proteins. HULSE/LAING: PROTEINS OF TRITICALE 13

TABLE 3. Average amino acid content (milligram per gram total nitrogen), (ranges in parentheses) of wheat, wheat products, rye and triticale, determined by column chromatography (data from Food and Agriculture Organization l 970b).

Wheat flour (70-80% Rye Triticale Wheat Wheat Wheat extraction whole (one sample Triticale• Amino acid meal germ bran rate) meal only) (1972-73)

Isoleucine 204 225 209 228 219 239 187 (188-214) (210--252) (193-226) (200--272) (200--242) Leu cine 417 433 415 440 385 402 450 (371-450) (408-490) (400--432) (376-643) (361-406) Lysine 179 407 270 130 212 192 196 (131-249) (369-517) (257-312) (108-194) (151-281) Methionine 94 122 102 91 91 87 60 (63-156) (111-164) (89-127) (63-122) (59-181) Cystine 159 130 168 159 119 79 (111-212) (110--172) (145-189) (123-208) (85-156) Phenylalanine 282 257 263 304 276 286 286 (234--338) (239-283) (255-279) (263-346) (250--300) Tyrosine 187 194 197 145 120 187 195 (86-225) (185-219) (178-211) (74--218) (76-175) Threonine 183 265 223 168 209 169 196 (148-222) (253-294) (164--254) (143-211) (191-231) Tryptophanb 68 66 80 67 46 63 (51-136) (56-78) (85-113) (65-70) (34--88) Valine 276 314 315 258 297 274 242 (228-325) (293-342) (233-371) (221-312) (206-343) Arginine 288 513 490 221 286 286 382 (234--344) (471-528) (461-540) (162-337) (184--344) Histidine 143 180 195 130 138 140 133 (125-163) (165-210) (175-217) (114--151) (125-165) Alanine 226 410 347 192 266 227 258 (188-314) (373-430) (319-371) (160--250) (235-302) Aspartic Acid 308 576 513 257 447 297 416 (263-338) (537-631) (474--556) (217-337) (383-511) Glutamic acid 1866 1102 1282 2184 1511 1860 1528 (1581-2019) (954--1287) (1144--1433) (1512-2733) (1356-1676) Glycine 254 393 405 222 271 245 265 (188-275) (375-440) (375-412) (183-271) (250--302) Pro line 621 343 395 726 586 705 521 (550--736) (290-426) (371-450) (556-849) (517-738) Serine 287 303 304 294 270 233 250 (256-319) (262-359) (260--342) (251-377) (250--306) First limiting' amino acid Lysine Isoleucine Isoleucine Lysine Tryptophan Second limiting' amino acid Isoleucine Tryptophan Lysine Threonine Isoleucine

"Calculated by the authors from data provided by Dr. E. Villegas and representing the means of three advanced triticale lines (6387a, l 1453a and l 142a) (tryptophan 2 samples only) produced at CIMMYT in 1972-73. bMicrobiological assay. 'The first and second limiting amino acids are as stated in the FAO publication. 14 MONOGRAPH IDRC-021e

Perhaps the most important general conclusions version factors for wheat (N x 5.61) and rye to be drawn from McLaughlan and Campbell's (N x 5.64) somewhat lower than those recom­ Chapter 2 is the urgent need for: mended by WHO (see Table 5). Tkatchuk (a) an internationally standardized and accepted recommends a factor of N x 5.76 for triticale. methodology by which to evaluate the biological A universally accepted convention of expressing quality of cereal and legume proteins; the amounts of individual amino acids present is (b) a clear differentiation between (i) approxi­ also needed. Different authors quote, for example, mate methods proposed for early identification lysine in terms of (i) % of protein (ii) % of total of promising plant materials, and (ii) more dry matter (iii) gram (or mg) per gram N (iv) precise biological methods by which to make a gram per 16 g N (v) micromoles per gram N final and definitive evaluation; (vi) micromoles per gram protein. (c) a standardized terminology and universal Convention (iv), g/16 g N, assumes a constant system of recording, computing and evaluating protein conversion factor of N x 6.25 for all foods analytical results related to protein content and (6.25 x 16 = 100). For cereals, however, a con­ quality, and amino acid composition; version factor between 5.7 and 5.8 is more (d) improved micro and preferably non-destruc­ appropriate hence lysine should be expressed as tive methods of analysis for protein and essential g/17.25 or g/17.5 g N. This is obviously a clumsy amino acids in early selections of cereal grains convention and therefore g (or mg) lysine/g N is and legumes, where only very small quantities of to be preferred. seed are available. McLaughlan and Campbell refer to several Equally important is the need for a much other methods requiring universal standardization closer working relationship in the future than in including, for example, the method of protein the past between the plant scientist and the hydrolysis before amino acid determination. This nutrition scientist and, also, between the food need for a standardized methodology is referred scientist and the nutrition scientist. While it is as a matter of urgency to the International Union fully recognized that the currently available of Pure and Applied Chemistry, the International methodology is inadequate and has many short­ Union of Nutrition Sciences, and the United comings, little is to be gained by introducing new, Nations Protein Advisory Group. seemingly more convenient, methods, if these new The need for micro non-destructive methods of methods are inaccurate, indiscriminating and ir­ analysis has been repeatedly urged by plant relevant to the analytical purpose intended. breeders seeking to develop cereals and legumes of The need for international standardization of improved nutritional quality. The plant breeder both methodology and terminology will be ap­ has available only a very small quantity of material parent to anyone who studies in depth the from his early lines, consequently methods of literature reviewed. Some authors see fit to analysis and selection which call for only a few mention neither the methods employed nor the seeds are a primary requisite. The methods will basis upon which their results are expressed. It be of considerable merit if they can be applied need hardly be emphasized that protein deter­ without damage to the seed's viability. mined by N x 6.25 and expressed on a dry weight This point again emphasizes the need for the basis will give a much higher result than N x 5.7 closest possible working relation between the at 14% moisture. For example let us assume, on plant scientist and the nutritional biochemist a dry weight basis, that N, by Kjeldahl, equals through the whole sequence of new varietal 2.50: development. A deeper and continuing under­ N x 6.25 (at zero moisture)= 15.62% protein standing of the plant breeder's difficulties might N x 5.70 (at zero moisture)= 14.25% protein serve to obviate some of the unreasonable requests N x 5.70 (at 14% moisture)= 12.00% protein made of him by nutritional scientists who, in some The highest result (15.62%) is 30% higher than the instances, do not appear to appreciate the com­ lowest (12.00%). plexity of changing the biochemical characteristics There appears to be some measure of disagree­ of a plant without prejudice to its other essential ment, even among knowledgeable analysts, con­ qualities. At the same time, a close cooperation cerning appropriate nitrogen conversion factors. would discourage plant breeders, motivated by a For example, Tkatchuk (1969) proposes con- justifiable sense of frustration, from adopting HULSE/LAING: PROTEINS OF TRITICALE 15 analytical and biological methods of selection suitability for use of the cereal or legume variety which, though economic and convenient, may developed. Wherever possible the final definitive prove misleading and in the long run counter­ evaluation should be made using the animal for productive. Much scientific effort may well be which the cereal is intended. wasted if the nutritional biochemist waits until McLaughlan and Campbell review several the final stages of varietal development before methods of determining protein nitrogen and becoming actively interested in plant breeding specify the conversion factors appropriate to programs. different plant protein sources. They discuss amino McLaughlan and Campbell review both acid analysis by chemical and microbiological chemical and biological methods of evaluation methods, the calculation of Chemical Score and pointing out the virtues and deficiencies of each the means of identifying limiting amino acids. method discussed. They describe the underlying They make reference to amino acid availability, principles and the factors which affect alternative pointing out that some of the amino acids recorded methods of biological evaluation including, among by ion exchange chromatography may be "un­ others, Protein Efficiency Ratio (PER), Net Pro­ available" nutritionally when the protein is fed to tein Utilization (NPU), Net Protein Ratio (NPR) an animal. and Biological Value (BY). They emphasize that The biological methods of protein evaluation the final evaluation of a cereal or legume must be described include those which depend upon body by a biological (animal feeding) method, at more weight gain and those which depend upon nitrogen than one level of protein intake, as described in retention in test animals. The slope assay referred their discussion of the Growth Slope assay method. to in Chapter 2 depends upon body weight gain; This point is further emphasized by Scrimshaw the slope assay described by Scrimshaw and Young and Young (1973) who state that in well-nourished (1973) depends upon carcass nitrogen retention. It young men the utilization of wheat gluten nitrogen would appear that the controversy among nutri­ is relatively high at low levels of intake but falls tional biochemists and physiologists concerning off at higher levels even though these higher levels the relative merits of biological methods based are still within the body nitrogen sub-maintenance upon weight gain versus those based upon range. They refer to other studies of wheat gluten nitrogen retention may continue for some time. utilization which support the conclusion that, for This clearly emphasizes the need for an inter­ proteins limiting in lysine, the efficiency of nitro­ nationally recognized body such as the Protein gen utilization drops markedly with increased Advisory Group of the United Nations, in con­ protein intake well before requirement levels are sultation with the appropriate international reached. Scrimshaw and Young state "It is clear scientific unions, to recommend and publish, for that more information about the ability of a the immediate guidance of plant breeders, accept­ protein to meet protein needs is obtained from the able and applicable methods of biological selection slope ratio approach than· from the assay of a and evaluation. protein at any single level of intake." They go At the conclusion of Chapter 2, McLaughlan on to state that while it may not be necessary to and Campbell, recognizing the plant breeders' abandon entirely single protein level biological need, present a series of recommended procedures assay methods, the differences between the nutri­ by which to determine the biological value of tive values of proteins of high and low quality are cereal and legume proteins. Briefly, where small more clearly demonstrated with the slope ratio samples of grain are available from early lines, method. they recommend the following determinations: The points made by McLaughlan and Campbell (i) Total Kjeldahl nitrogen. and by Scrimshaw and Young emphasize the (ii) Amino acid content by ion exchange plant breeder's need for several standardized chromatography or amino acid auto­ methods: first, those which can be used for early analyzer. sorting of new lines; second, methods which are more precise and comprehensive to be used as (iii) The limiting amino acids, based upon sufficient plant material becomes available; and FAO/WHO reference patterns. finally, biological methods by which to determine (iv) Net Protein Ratio assay using 5 rats for definitively the overall biological quality and IO days. 16 MONOGRAPH IDRC-02Ie

For a more complete biological evaluation where In reviewing the wheat and rye literature, emphasis larger samples are available, they recommend a has been placed upon those protein factors which Growth Slope assay at no less than three levels of influence biological value in human nutrition protein intake. since the literature relating to varietal and en­ vironmental influence upon nitrogen content, The remainder of the text covered in the review particularly in wheat, and its use in feed and forage, falls into two broad categories: is so vast as to be virtually unmanageable in a A. Research by plant scientists. review of this kind. Consequently, this aspect of B. Research by food and nutrition scientists. the text has been confined to a few basic references. Even a cursory review of the subsequent text will reveal a lack of uniformity in content, con­ GENETIC INFLUENCE siderably more having been published on certain The chapter begins with a brief discussion of aspects of the subject than others. Since most of the complexities of plant breeding and genetic the research published has been undertaken in influence as they relate to cereal protein com­ North America and the developed nations of position. Since the precise composition of the Europe, the literature heavily emphasizes bread embryo protein is vital to the plant's survival, any wheats. Furthermore, greater attention has been intended genetic manipulation must concentrate given to those factors which influence economic upon the storage proteins of the endosperm. The value (including yield capability of the grain, and discovery in maize of certain genetic mutations water absorption and "baking strength" of the which depress the prolamin content of the protein derived flours), than to nutritional values of wheat and thereby increase the proportion of lysine and and rye proteins. This review deals almost ex­ other essential amino acids, together with the clusively with the nutritional aspects of the advent of new processing technologies which are subject. less dependent upon high gluten contents, en­ Though plant scientists have striven for many courage the continued search for high lysine years to increase yield potential and total protein (prolamin depressant) genetic mutations in wheat content, since both are factors of economic im­ and rye. portance, until very recently they have shown little Protein content and composition vary among interest in amino acid composition as it relates to different fractions of the cereal grain seed. In the biological value of the cereal grains. The food selecting for high protein genotypes it would scientists and cereal technologists have occupied appear more meaningful to select on the basis of themselves largely with the functional properties protein content per seed rather than protein as of cereal grains and with the development of percent dry matter. Protein as percent dry matter labour saving and more profitable methods of is influenced by seed weight and the relative breadmaking and other processing technologies. proportions of the various seed fractions present. Those nutritionists who have explored the bio­ These in turn are influenced by environment and logical value of cereals appear to have been more agronomic conditions. interested in fortification and supplementation The literature reviewed clearly indicates that than in the possible influence of genetic back­ while the potential to deposit a high proportion ground, agronomic and environmental history of protein nitrogen in the endosperm of triticale, upon nutritional value. wheat and rye is controlled genetically, the amount actually deposited, (i.e. whether the grain achieves Varietal and Environmental Factors its full potential for protein production) is greatly Chapter 3 describes the influence of genetic, influenced by environment and agronomic varietal, environmental and agronomic factors management. Because of these strong environ­ upon the biological value of the protein oftriticale mental and agronomic influences, any potentially and its parents. Both wheat and rye are used as superior genetic material must be widely tested food and feed grains and as forage crops, and it is and confirmed before a final judgment is pro­ probable that triticale will be similarly used. The nounced. review specifically related to triticale attempts to While there appears to be some difference of cover comprehensively its protein content and opinion, there is evidence to indicate that the biological value for humans and other animals. protein of triticale possesses its own unique HULSE/LAING: PROTEINS OF TRITICALE 17 characteristics and its spectrum cannot be regarded At the University of Manitoba, triticale was as a simple addition of the protein spectra of its compared with wheat and several other cereal two parents. Nevertheless, all relevant factors grains in feeding tests with rats, laboratory mice being equal, the analysable protein nitrogen con­ and the meadow vole (Micro/us pennsylranicus). tent of triticale appears to fall between the pro­ Though the standard error was lowest with the tein contents of its two immediate parents. This rats, both rats and mice clearly indicated the appears to be also true of the proportion of superior biological value of triticale over wheat essential amino acids present, the lysine content at two protein levels. The results from the voles of triticale being generally higher than in wheat appeared totally unreliable, the standard error but lower than in rye. being of the order of 50% of the means. Further­ One of the most interesting observations comes more, the voles rated triticale and other cereal from Riley and Ewart (1970) who report a sig­ grains biologically superior to casein, the standard nificant interchromosomal interaction when dif­ protein of comparison. ferent pairs of rye chromosomes were added separately and in turn to a wheat genotype. The TRITICALE FED TO FARM ANIMALS content of several amino acids in triticale appeared In trials with chicks, the results are variable to be influenced by interchromosomal interaction among several reported experiments, the vari­ whereas others were not. The chromosomes in ability being probably attributable in some degree homoeologous group 5 appeared to be of partic­ tci the varying composition and proportion of ular interest in promoting a higher lysine content. dietary ingredients other than triticale and the cereals with which it was being compared. In LYSINE IN TRITICALE general, however, it appeared that triticale was at As is detailed more precisely for wheat, the least equal to, if not better than, wheat and that lysine content of triticale expressed as percent of lysine was the first limiting and methionine the total protein is generally inversely correlated with second limiting amino acid. The difficulty of protein as percent dry matter. Lysine as percent evaluation arose, in some instances, since soya total dry matter is positively correlated with protein was used to bring all test diets to the same protein expressed on the same basis. nitrogen level. It is probable in such instances From the outset of the Triticale Project, triticale that any differences between triticale and other lines have demonstrated lysine contents superior cereals are obscured by differences in soya protein to wheat. This superiority is evident again in the content. Also, as a general comment on animal preliminary screening of CIMMYT's 1972/73 feeding trials, too little attention appears to have selections where lysine (as percent of protein) been given, or at least recorded, concerning the reportedly ranged in sixteen samples from 3.72 characteristics of the triticale used, particularly (13.0%) to 4.35 (13.0%), the figures in parenthesis with regard to the size and condition of the being protein (Kjeldahl N x 5. 7) contents. Villegas kernels, and to what degree the triticale was free (Zillinsky 1973) reports advanced (1972-73) triti­ from ergot contamination. From the data derived cale lines at CIMMYT equal in lysine content but 0 from early triticale lines it would appear that the significantly superior in protein content to opaque- utilization of triticale and wheat protein by chicks 2 maize. are roughly equivalent. In general, and in the BIOASSA YS OF TRITICALE light of some recent data, triticale, if free from The biological value of triticale vanelies has ergot, appears to be a satisfactory cereal for in­ been assessed in a number of animals. In human clusion in the diets of broilers, laying hens and feeding experiments, in terms of nitrogen reten­ turkey poults. tion, triticale appeared superior to wheat at two The earlier results of hog feeding trials suggest levels of protein intake. Lysine was established that triticale is equal to barley for heavier animals as being the first limiting amino acid. Lysine was but inferior in the diet of lighter pigs. Some also first limiting in rat feeding experiments in authors report reduced palatability and loss of which, as assessed by PERs, triticale appeared appetite when hogs were fed large proportions of superior to wheat. On the other hand, in labora­ triticale. Again, several authors do not indicate tory mice, methionine was reported as first whether the triticale used was clean or infected. limiting both for triticale and for wheat. Where ergot infection was clearly demonstrated, 18 MONOGRAPH IORC-02le marked reductions in feed intake and weight gain increased and the average protein content has were recorded. Some authors report triticale as noticeably declined. During the late 1960s protein being satisfactory as a feed for growing-finishing contents (N x 5.7) of hexaploid triticale lines swine but one author recommends that triticale grown at CIMMYT ranged from 11.7% to 22.5% be restricted to 25% of the ration for growing pigs, with an average of 17.5%. These early lines to 50% of the ration for finishing pigs, and that it included a large proportion of shrivelled kernels. be not recommended in hog starter rations or for The advanced hexaploid lines grown at CIMMYT breeding stock. Others report triticale, when fed during 1972 ranged in protein from 10.9% to as 95% of the diet of hogs, as being equal to wheat 19.1% with an average of 13.4%. It should be and better than barley. There appears general noted, however, that in the earlier lines the lysine agreement that lysine is the first limiting amino content (percent of protein) ranged from 2.5% to acid in triticale for swine. 3. 7% with an average of 3.2%. Samples from 1972 Triticale when fed to dairy calves resulted in trials ranged in lysine from 2.6% to 3.9% with an small reductions in feed intake and weight gain average of 3.4% (Zillinsky 1973). As stated above, compared with barley-urea and barley-soya. The preliminary screening of 1972-73 materials indi­ relative feed efficiencies were reported as not cates a continued improvement in lysine content. significantly different. Again, no reference was While there is still much to be learned about made to possible ergot contamination. Several the precise influence of genetic background upon trials with steers demonstrated lower feed con­ the protein content and composition of triticale, sumption with triticale though the protein digesti­ there is adequate evidence to predict with con­ bility and feed efficiency of triticale appeared fidence that triticale will take its place as a high superior to both wheat and sorghum. Feed intake yielding, high protein cereal crop of comparatively improved when the triticale was steam-rolled. superior amino acid composition and of satis­ Some authors reported a higher incidence of factory digestibility and feed efficiency. liver abscesses in steers fed triticale. Steam-rolled triticale appeared equal to steam-rolled barley for WHEAT PROTEIN The literature on wheat is so vast that neither lactating Holsteins. In general, triticale was time nor space permit a comprehensive review. reported at least equal to wheat in sheep rations. Consequently, the text consists of a review of It must be borne in mind that many of the animal feeding studies reviewed are based upon early certain important publications drawn from as many geographical areas as possible. triticale lines most of which originated from From the extensive literature relating to wheat, breeding trials and yield nurseries. Furthermore certain clearly defined patterns and relationships there is a marked paucity of information con­ cerning the overall quality and characteristics of are evident. For example, among wheats con­ taining protein content (N x 5. 7) up to about 14%, the triticales tested. It is therefore desirable that lysine expressed as percent protein is significantly many of the animal feeding studies be repeated negatively correlated with protein expressed as using advanced genotypes of known history and percent dry matter. Above 14% protein the nega­ identification. tive correlation is not significant and disappears ADV AN CED TRITICALE LINES entirely above 16% protein content. Lysine ex­ Considerable progress has been made during pressed as percent of total dry matter in the grain the recent past in increasing triticale yields, in is positively correlated with grain protein content. reducing the incidence of grain shrivelling, and in The highest lysine value reported from the World improving kernel characteristics. Yields oftriticale Wheat Collection is about 4.3% of protein. in excess of 8000 kg/ha, significantly higher than The total protein nitrogen deposited in the the highest yielding bread wheats grown under endosperm is greatly influenced by environmental comparable conditions, are reported from and agronomic conditions, therefore, as stated CIMMYT. As reported above, the CIMMYT above, it is necessary to grow any apparently advanced lines continue to demonstrate com­ "high protein" wheats over several years in a paratively high lysine contents. variety of locations in order to establish a truly However as kernel characteristics have im­ genetic high protein capability. Threonine, proved, the ratio of endosperm to bran has generally considered the second limiting amino HULSE/LAING: PROTEINS OF TRITICALE. 19 acid in wheat, and leucine, appear to be positively The response of rye to nitrogen fertilizer correlated with lysine. appears to parallel that of wheat, in that grain Within the World Wheat Collection, protein nitrogen increases with the addition of nitrogenous contents on a dry weight basis range from 6 to fertilizer, the main increase being in the pro lam in 23% and lysine as percent of protein from 2.2 to fraction. In general, the literature indicates higher 4.3%. While no high lysine gene in wheat com­ lysine values, expressed as percent of protein, parable to the mutations found in maize has yet in rye than in wheat, the highest encountered in been discovered, it is important to note that in this review being 5.3%. terms of essential amino acids expressed as per­ cent of total dry weight matter, high protein Nutritional Inhibitors and Toxic Factors wheats provide more of all essential amino acids Chapter 4 describes toxic substances and than low protein wheats. nutritional inhibitors naturally present and ac­ Neither protein nor lysine contents in wheat quired in the cereal grains under discussion. In appear influenced by the size of the kernel pro­ general, too little scientific attention has been vided it is plump, healthy and adequately filled. given to such substances to permit a clear under­ Shrivelled and deformed kernels with a high bran standing of the mechanism of their physiological and germ to endosperm ratio tend to give high action in humans and other animals or to the protein and high lysine analyses. overall importance of their influence upon human The influence of environmental conditions upon and animal nutrition. wheat protein nitrogen content is well established. Resorcinol derivatives have been reported in Significant varietal differences in response to nitro­ rye, triticale and wheat, generally being found in gen fertilizer, indicating genetic control of total highest concentration in rye and lowest in wheat. protein content, have been recorded in many They appear to be concentrated in the pericarp publications. In general the response to increased and therefore appear in higher proportion in nitrogen fertilizer is an increase in the prolamin shrivelled than in well-formed, plump grains. fraction. Several authors have reported a linear There is some, but by no means complete, evi­ relation, given the necessary genetic potential, dence that their presence is influenced both between protein nitrogen content and the rate of genetically and environmentally. nitrogen fertilizer applied. Though increase in The symptoms of resorcinol action appear to be fertilizer increases the proportions of proline and a depression of appetite and a lowered food intake. glutamic acid which predominate in the prolamin Resorcinols have been shown to be present in both fraction, neither environment in general nor rye bran oil and wheat bran oil and to be sig­ fertilizer application in particular appears to in­ nificantly reduced in activity during baking and fluence the relative proportions of the essential steam-rolling. In Chapter 5, which deals with amino acids present in wheat protein. processing, it is suggested that some attention Some evidence is presented that certain herbi­ might be given to removing the pericarp by con­ cides lead to an increase in total protein nitrogen trolled abrasion milling. This might also serve but they do not appear to affect essential amino to remove undesirable microorganisms attached acid composition. to the outer grain surface. Phytic acid and its derivatives are not known to RYE PROTEIN be of importance in protein metabolism but, since Several authors have described how the bio­ phytic acid is associated with the higher protein logical value of rye protein is superior to wheat fractions of wheat and rye, it deserves mention. protein. Since rye is of minor significance in world Phytic acid forms insoluble salts with the dietarily dietary patterns compared with wheat, rice and important elements calcium, iron, magnesium and maize, comparatively little research has been zinc, thus rendering them unavailable to the devoted to improving its agronomic and biological organism. Phytic acid is enzymatically destroyed properties. A more thorough examination of during dough fermentation and during certain varietal influence upon protein content and stages of cooking and baking. Its importance quality in rye might well prove fruitful in breeding should not be overlooked in those less developed nutritionally superior triticale varieties. countries where the phytic acid present in high 20 MONOGRAPH IDRC-021e extraction cereal flours may seriously affect the their physiological action is influenced by the absorption of calcium, iron, magnesium and animal species, its size, age and general health and zinc present in infant diets. by the total quantity of food consumed. Where Trypsin inhibitors are present in a wide variety tolerance limits are recommended it is essential of edible plant seeds including wheat, rye and that the required methods of sampling, inspection probably triticale. While their action may be of and analysis also be precisely defined. significance in raw cereals fed to animals, it is not unlikely that they are of minor importance in Processing and Supplementation cooked cereals. In Chapter 5, which deals with the technologies Factors other than resorcinol derivatives which of processing and supplementation, very little is depress growth have been reported present in to be found on how the biological value of rye. Their action appears to be offset by anti­ triticale is influenced by milling, baking, other biotics and by autoclaving in water or mild acid. processing, or by supplementation with additional It is possible that these unknown substances may protein sources. On the other hand, there is a stimulate the growth of undesirable intestinal substantial literature relating to the biological microflora but their nature and mode of action value of processed wheat and rye products, with have not been positively identified. There is some and without nitrogenous supplements, much of evidence to suggest that, unlike the resorcinols, which is relevant and applicable to triticale. these unknown growth depressants are located in Since virtually all cereals are processed before greater concentration in the endosperm than in being eaten, and since the supplementation of the bran. cereal flours when processed into bread, other Ergot is one of the most serious contaminants baked products or alimentary pastes, is neither to which triticale appears susceptible. This sus­ novel nor technically difficult, the influence of ceptibility is inherited from its rye parent and both processing and supplementation is believed also from the tetraploid T. durum from which to be relevant and germane to this review. most existing triticales are derived and which is Once again, the task would be less difficult if all more susceptible to ergot than the hexaploid of the authors reported could have employed a T. aestirnm. Ergot is considerably influenced by standardized methodology and terminology and environment and is virtually unknown in some of shown a greater attention to detail in describing the regions where triticale might be grown. In both the cereals used and the methods by which several reported instances, both feed intake and they were processed and analyzed. One might weight gain have been inversely related to the suggest that if the research is worth doing, it is concentration of ergot. worth the effort to determine the nature and A considerable volume ofliterature has appeared history of the cereal grains and other raw materials in recent years concerning aflatoxins. Apart from used and to specify the conditions of processing. the serious acute and chronic toxicity symptoms The importance of describing the physical attributable to the presence of aflatoxins, the characteristics of cereal grains used in nutritional presence in grain of the mould fungi responsible studies, and in particular whether they are plump has been shown to reduce significantly the pro­ or shrivelled has already been emphasized. portion of total amino acids and of certain essential amino acids originally present. Other MILLING OF TRITICALE mould fungi may cause a loss in protein quality There is insufficient data on which to base any in cereal grains and grain legumes by selectively definitive conclusions concerning the milling destroying some essential amino acids. qualities of triticale. Though triticale appears to In addition to the mycotoxins reviewed in respond satisfactorily to conventional laboratory Chapter 4 there may be others as yet unidentified. milling technologies, the comparatively high pro­ A recent verbal communication to the authors portion of inadequately filled kernels among early indicates that some species of the genus Fusarium lines has tended to give lower milling yields. known to infect triticale, are able to produce Villegas (Zillinsky 1973) refers to milling tests mycotoxins. using a laboratory mill in which flour yields ranged Tolerance limits for ergot and mycotoxins are from 51.7% to 59.0%, compared with the control extremely difficult to prescribe since the extent of wheat !NIA 66 which yielded 83.7% flour. HULSE/LAING. PROTEINS OF TRITICALE 21

In the light of the considerable improvement The digestibility of uncooked wheat and rye flours evident in the kernel characteristics of advanced decreases with increasing extraction rate. Con­ triticale lines it is recommended that more sequently, the biological superiority of high attention now be given to milling and other extraction versus low extraction endosperm flours technological characteristics. Both CIMMYT and is somewhat offset by reduced digestibility. In rye, the University of Manitoba are examining the as in wheat, the percentage protein content and technological and functional properties oftriticale. the lysine both as percent protein and as percent In addition, in cooperation with the International total dry matter, increase with extraction rate. Union of Food Science and Technology, an inter­ Similar trends are evident in triticale. To what national working group has been formed to ex­ degree the improved protein pattern of higher plore how triticale might best be processed and extractions flours is offset by decreased digestibility, used in cereal foods in various countries through­ appetite-depressant and other inhibiting factors out the world. deserves serious attention among the more ad­ It is recommended in the text that triticale be vanced triticale lines. The nutritional value as subjected to abrasion milling to remove the seed influenced by processing conditions needs to be coats of the pericarp and thus reduce the resorcinol examined in the diets of a variety of animals. content together, hopefully, with most of the un­ desirable microorganisms present on the surface BAKING WITH TRITICALE of the grains. A few authors have reported that satisfactory Of particular interest are a few results obtained bread can be baked from flours containing high from air classification experiments in which proportions of triticale flour though none of the triticale lines displayed significantly higher protein research reported gives any account of the bio­ shifts than either wheat or rye. No evidence has logical value of the bread produced. Since most been presented to suggest that the amino acid of the triticales available are hexaploid and composition of high protein, air classified flour is derived from a durum wheat parentage, in addition essentially different from triticale flours processed to which alpha-amylase activity tends to be higher by conventional milling. in triticale than in wheat, it is not to be expected that triticale will, by conventional fermentation WHEAT AND RYE FLOURS methods, produce bread equivalent to flour from Flours from low protein soft wheats tend to be bread wheats. Nevertheless some results suggest higher in lysine, arginine and several other that by adopting new breadmaking technologies, essential amino acids than high protein strong including those based upon mechanical develop­ wheat flours. The lysine content of wheat flours, ment, satisfactory bread is possible from triticale and probably triticale flours, decreases with flours. decreasing extraction rate since the inner bran and, Villegas (Zillinsky 1973) reports that acceptable in particular, the aleurone layers are richer both tortillas and cha pa tis can be made from I 00% in total protein and lysine than the inner endo­ triticale flour. A private communication indicates sperm. Lysine is the first limiting amino acid in that triticale flour has been found acceptable as a both wholemeal and white (endosperm) flours and 75% replacement for teff and other grains in in the unsupplemented bread made from them. traditional Ethiopian cereal foods. From the point of view of protein content and Though little is published on the processing of biological quality, 85% extraction is significantly triticale, the effect of baking upon its nutritional superior to 75% extraction flour. value would not be expected to differ greatly from The addition of bran to white flours increases rye and wheat. Baking of both wheat and rye flours body weight gain and biological value in rats reduces the available lysine content, the loss, all though the addition of bran alone has not, as other things being equal, being a function of time judged by PERs, been shown to equal the optimum and temperature. In bread, therefore, the greatest supplementation of white flour with synthetic reduction occurs in the crust, and lysine losses are lysine and threonine. highest in bread in which the crust to crumb ratio While there is some variation among other is highest. Rye crisp breads, for example, suffer essential amino acids, the availability of lysine in greater damage to protein biological value during wheat and milled flour are not greatly different. baking than do "soft" rye breads. Bread baked by 22 MONOGRAPH IDRC-021e microwave heating is biologically superior to oven portional to the L-lysine added to wheat flour up to baked bread in that the time of microwave baking a level of between 0.2 and 0.25 g lysine per 100 g is extremely short and no crust is formed. wheat flour at which point lysine ceases to be the According to several authors, there is no first limiting amino acid. Several papers report the significant decline in protein value when high significant loss of added lysine during baking, extraction wheat flour (atta) is baked into cha patis. probably attributable to reactions with reducing It is suggested that any lysine loss is compensated sugars present. The unavailability of added lysine by improved palatability and digestibility. It may in general increased with increase in baking time. also be that chapati doughs are proportionally In infants fed wheat semolina some benefit of lower in reducing sugars than fermented bread adding lysine and potassium together was demon­ doughs in which maltose is known to be formed. strated. During baking, reducing sugars react with, and When lysine and threonine, the second limiting render, lysine partially unavailable. Puris, which amino acid, were added to wheat flour to the point are fried in oil, were lower in PER than chapatis of maximum nutritional efficiency, the resultant baked from the same atta. wheat bread protein was rated "first class", that is, essentially nutritionally complete. Added lysine OTHER PROCESSED CEREAL FOODS improves the nutritional value of most wheat Both the available lysine and biological value products including bread of various western and of pastas (macaroni, spaghetti, etc.) decline with oriental kinds, alimentary pastes (macaroni, etc.), increasing drying temperature. In breakfast cereals chapatis, and biscuits (cookies). The beneficial the biological value does not appear damaged by effect of added lysine appears most marked in cooking in water but may be lowered by toasting, the rate of weight gain in rats but improved puffing and high temperature extrusion. There is nitrogen retention with added lysine is also evidence that steam or hot water cooking of wheat demonstrable. It is also reported, at least over followed by drying, as in the production ofbulgur, short periods of time, that added methionine significantly increases the metabolizable energy improved an all-cereal diet fed to adults. Addi­ and may improve the biological value of the tions of methionine plus lysine were reported to be protein, though some authors report no improve­ beneficial in certain wheat diets. ment in protein quality between bulgur and the The need for further research into the role of original wheat. the "non-essential" amino acids present in cereals Fermentation of cooked wheat and mixtures of is stressed by several authors. Some results indi­ wheat and soybeans by Rhizopus organisms found cate that added lysine gives rise to a higher food in Indonesian tempeh, significantly increased the intake by rats. The reliability of results based protein value, fermentation of wheat flour plus upon weight gain in young rats fed amino acid soya flour giving biological values almost equal supplemented cereal diets has been questioned by to casein. one recognized authority. Cereal Proteins Supplementation with cereal PROTEIN SUPPLEMENTATION proteins has, until recently, focussed largely upon The review of supplementation deals only with added gluten, produced by washing out the starch nutritional implications and covers supplementa­ from a wheat flour dough. While added gluten tion by synthetic amino acids, cereal protein increases total protein, it does not significantly concentrates, egg and milk proteins, legume and improve nutritionally the balance of amino acids oilseed proteins, microbial ("single cell") proteins in any of the cereals under discussion. The and proteins derived from fish. The technological nutritional benefit of adding supplements derived consequences of adding various protein supple­ from wheat germ and wheat bran is reported by ments have recently been reviewed elsewhere several authors. (Hulse, in press). The methods and benefits of producing and Amino Acids Since lysine is demonstrably, using wheat protein concentrates are described. for humans, the first limiting amino acid in wheat, The most recent concentration processes entail the rye and triticale flours, most of the literature fine grinding and screening of certain selected mill relating to synthetic amino acids focusses upon feed fractions which are demonstrably higher in lysine, usually administered as L-lysine hydro­ both total protein nitrogen and lysine content chloride. Weight gain in rats is roughly pro- than the ground endosperm. The benefit to rate of HULSE/LAING: PROTEINS OF TRITICALE 23 weight gain and nitrogen retention of such wheat centration of nucleic acids present, excessive protein concentrates is reported by several authors. ingestion of which by monogastric animals leads Milk Proteins Apart from the increase in to undesirable physiological effects, including total protein nitrogen, the beneficial effect of kidney stones. The need for more research into the adding egg or milk proteins appears to be pro­ effects of high intakes of nucleic acids from portional to the resultant increase in lysine. It is microbial protein has been recommended by claimed that, as judged by PER, the addition of several authorities (United Nations 1968). While 6% nonfat dry milk (skim milk powder) is roughly there are methods available for removing nucleic equivalent to the addition of 0.17% lysine. One acids the resultant increase in cost might prove author reports that 18% of skim milk powder is prohibitive for developing countries. necessary to achieve a biological value equivalent Fish Proteins A good deal of attention has to the addition of L-lysine and L-threonine added been given to proteins derived from fish, partic­ to the point at which each cease to be limiting. ularly fish protein concentrate produced by extrac­ Significant losses in PER were reported when tion with isopropanol or other organic solvents. wheat flour supplemented with milk was baked Unfortunately, all of the literature does not into bread. Skim milk powder significantly im­ clearly differentiate among the various levels of proved the nutritional value of alimentary pastes, refinement possible in producing fish protein though there is evidence that some leaching of concentrates. In general, fish protein is a good soluble proteins may take place during cooking source of lysine and any significant additions to in hot water. cereal products improves the biological value in Legume Proteins For the less developed proportion to the amount of lysine added when world, the food legume and oilseed proteins offer the diets are judged isonitrogenously. The eco­ the most attractive source of supplementation. nomic feasibility of producing fish protein con­ Since most of the literature reported originated in centrates which are sufficiently refined to be North America, soya flour is dealt with at greater acceptable in odour and flavour when blended length than any other legume protein source. It is with cereal products has yet to be demonstrated. worth emphasizing that "soya flour" is by no Benefits and Cost of Supplementation In means a universally homogeneous material and some of the literature reviewed it appears to be that its biological value is significantly influenced assumed that any and all forms of protein supple­ by processing conditions. Some evidence indicates mentation are desirable. It must be emphasized that within limits the biological value of soya that an ill-informed approach to protein supple­ protein improves with heat treatment, possibly mentation, particularly with synthetic amino as a result of the destruction of trypsin inhibitors acids, may give rise to undesirable protein nitrogen and other antinutrient factors. Soybean isolates imbalances, particularly ifthe supplemented cereal produced by alkaline extraction followed by acid provides most of the protein calories consumed. or heat precipitation are of significantly lower The problems of amino acid imbalance have been biological value than defatted heat-treated soya discussed by Bender (1965), Carpenter and de flours. Muelenaere (1965), Harper (1969), Harper and Though the evidence is much less weighty there Rogers (1965), Kies and Fox (1972), Sanahuja are clear indications that other oilseed and legume (1971 ), Muramatsu et al. (1972). However, where proteins can serve as useful supplements to cereal protein malnutrition is apparent or likely to occur, products. Defatted cottonseed flour and proteins appropriate supplementation of a predominantly derived from chick-pea, pigeon pea and other cereal diet with an inexpensive protein source is tropical legume flours have been shown to be to be recommended, particularly for young nutritionally beneficial. In most instances the children. benefit, apart from the increase in total protein In general, as judged by PER and other indices nitrogen, appears to be roughly proportional to based upon body weight gain, the nutritional the lysine added. benefit derived from supplementing processed Microbial Proteins Yeast and other micro­ wheat, rye or triticale with other protein sources organisms offer potentially useful sources of is proportional to the amount of lysine added protein supplementation. The level to which they until the point at which lysine ceases to be the can be usefully added appears to be limited, not so first limiting amino acid. McLaughlan and much by their amino acid content but by the con- Morrison (1960) report that in rat diets in which 24 MONOGRAPH IDRC-021e half the protein was supplied by bread and the TABLE 4. Effect on protein content of various supple­ other half by each in turn of(a) oatmeal, (b) whole ments to wheat flour. dried eggs, (c) dry beans, (d) casein, (e) cheese and (f) fish flour, there appeared an almost perfect (b) correlation (r = 0.99) between PER and total Approximate lysine content, the PERs increasing progressively (a) increase in from (a) to(/), unsupplemented bread being the Approximate protein lowest, bread plus fish flour the highest. quantity of adding necessary (a) to 100 g It might be concluded therefore that the sole to add wheat flour purpose of supplementation should be to add 0.25 g of 12% protein lysine until it ceases to be limiting and that this lysine content might be best achieved by adding L-lysine hydro­ (g) (%) chloride. It must be remembered, however, that many of the comparisons reviewed in Chapter 5 Egg 9.0 19 are based upon PERsdetermined in isonitrogenous (whole dried) Milk 9.5 17 diets. In practice, however, different protein (nonfat dried) supplements which contribute the same quantity Soya 8.0 23 of lysine will add significantly different amounts (defatted) of total protein nitrogen to the diets. Altschul Cottonseed 10.0 36 and Rosenfield (1970) state that 0.25% is the level (de fatted) at which lysine ceases to be the first limiting amino Peanut 15.0 40 acid in wheat flour. Table 4 gives (a) the approxi­ (defatted) mate quantities of various protein supplements Chick-pea flour 17.0 12 in grams per I 00 grams of flour necessary to pro­ Fish protein concentrate 3.0 18 Yeast 6.0 18 vide 0.25 grams of lysine and (b) the approximate (dried) percent increase in protein which would result if Wheat protein such additions were made to a cereal flour of 12% concentrate 24.0 17 protein. It is evident that all supplements other Lysine 0.25 0 than lysine will significantly increase the total protein content. protein, food calories and other nutrients. Further­ Furthermore, the cost of supplementation to more if, for example, the cereal flour costs I 0 the same level of lysine will vary greatly among cents per kilo and an added legume flour costs I 0 protein sources. Hulse (in press) estimated in cents per kilo there will be no increase in cost per 1971 that the ingredient cost of adding 0.25 lb kilo of the final supplemented mixture. Unless of lysine to I 00 lb wheat flour in Canada varied the cost per unit weight of the supplement is from nearly $7.00 for dried egg through $2.30 for equal to, or lower than, the cost of the cereal flour, skim milk powder, $0.56 for defatted soya to any degree of supplementation will inevitably $0.25 for L-lysine hydrocholoride. Obviously these raise the cost of the final product. costs will differ among countries and will change It is the authors' opinion that where necessary,· from year to year. At the moment of writing and as far as circumstances permit, the poor (October 1973) all of the relevant prices are nations would be best advised to study seriously markedly higher than in 1971. the means of supplementing cereals with flours Though L-lysine hydrochloride appears least derived from indigenous or locally adapted expensive it is available from only one country, legumes which generally are much cheaper than Japan. Continued use by another nation would most other protein supplements. It is the authors' therefore call for a continued expenditure of intention, in the near future, to prepare a review foreign currency. Many developing nations would of the nutritional value of certain important doubtless encounter technical and logistical tropical food legumes and the products derived problems in uniformly distributing and dispersing from them. The authors also propose to prepare a lysine throughout all of their home-produced review similar in scope to the present publication cereal flours. Nor does added lysine increase, as in which the biological values of the sorghums and do the other supplements, the total quantity of millets will be considered. " •• trt11cafe.1 w1ff soon add to tire 1"orfd '.1 loml prod11ctio11 poremiaf" ( Norman Borltmg J Chapter 2

METHODOLOGY FOR EVALUATION OF PLANT PROTEINS FOR HUMAN USE

J. M. MCLAUGHLAN AND J. A. CAMPBELL* Health Protection Branch Health and Welfare Canada Ottawa

Many workers, including a recent expert group should bear a close relationship to those of (United Nations 1971 ), have stressed the need for humans (Campbell and McLaughlan 1970; increased intake of good quality protein, partic­ Bressani et al. 1973). ularly by certain segments of the population in Because of the fact that the protein value of a developing countries. Probably the simplest way food depends primarily on the content of the to meet this need is to develop new high protein limiting amino acid, which may vary from food to varieties of crops presently grown in the country food, the problem of protein evaluation has been or region in question, so that problems of use and a difficult and controversial one. Thus, many acceptability are minimized. Plant breeders have methods have been recommended and used and accepted the challenge and have attempted to their relative merits and limitations discussed at improve not only the yield, but also the protein length (Campbell 1963; National Academy quality and protein content of staple cereal and of Sciences-National Research Council 1963; legume crops for human use (Protein Advisory McLaughlan and Campbell 1969; Porter and Group 1971). Rolls 1973). In any program ofcrop improvement, considera­ In the development of new cereal and legume tion must, of course, be given to the possible varieties, protein evaluation revolves largely impact of the program on the nutritional status around a consideration of the lysine, methionine, of the population of the country or area. Thus, it threonine and tryptophan content of proteins, may not constitute good nutritional policy to but may, however, be subject to particular replace legumes with cereals simply because high difficulties. Some of the problems involved have yielding cereals can be produced. (Protein Advi­ been reviewed in recent documents of the Protein sory Group I 972a). Such a course of action may Advisory Group (1971, I 972b, 1973). However, reduce, rather than increase, the intakes of some in view of the specific problems related to the essential amino acids. A very clear assessment must needs of plant breeders, it was considered neces­ therefore be made first of the nutritional needs of sary in a review such as this to include a section on the particular area or country involved and of the protein evaluation. It is therefore the purpose of possible impact of any plant breeding program on this chapter to present a critique of existing the nutritional status of the population. The methodology and to suggest appropriate pro­ program should then be set up to meet these needs. cedures for the evaluation of protein in cereal Protein quality may be defined as the capacity grains and legumes for human use. Other recent of a protein or a mixture of proteins to satisfy reviews are available (McLaughlan and Campbell the requirements of the consuming animal for 1969; Porter and Rolls 1973). essential amino acids. If the food under study is for human use, the requirements of the test animal Need for Methods *Present address (J.A.C.): Caribbean Food and Nutri­ The need for nutritional and food quality guide­ tion Institute, Box 140, Kingston 7, Jamaica. lines, including methods for protein evaluation, 27 28 MONOGRAPH IDRC-021e has been clearly pointed out (Protein Advisory TABLE 5. Factors used for converting nitrogen to Group 1971 ). The plant breeder is faced with the protein (World Health Organization 1973). fact that he must test small quantities of newly developed lines at the earliest possible opportunity Conversion for a variety of qualities including yield. agronomic Foodstuff factor characteristics and nutritive value for the intended end use be it human food or animal feed. Ideally. CEREALS Wheat methods are therefore needed which will be Whole meal or flour or bulgur 5. 83 applicable to individual seeds to evaluate their Flour, medium or low extraction 5. 70 possible nutritive value and the presence of toxic Macaroni, spaghetti, wheat pastes 5. 70 or harmful substances. While this may not yet be Bran 6.31 practicable. there is an urgent need for the develop­ Rice ment of methods applicable to very small samples­ Husked or brown (only hulls removed)} at least as screening methods. Home~pounded, undermilled, . 5 95 In the development and use of such screening parb01led methods. it is essential that their meaning and Milled, white limitations be known and recognized and that the Rye assays be properly standardized. Until appro­ Whole meal, dark flour priate methodology can be developed for small Flour, medium extraction samples. recognition must be given to the fact Flour, light, low extraction that larger samples may have to be used for the Barley 5.83 assessment of protein value and that such testing Whole seed, except hulls and groats may have to be conducted at a later stage of the Pearled, light or dark breeding program. It is also obvious that careful Oats consideration must be given to the priorities Oatmeal, rolled oats involved in testing. Thus. if one of the prime purposes is to increase protein value of a crop. PULSES AND SEEDS protein testing should be initiated at an early Groundnuts 5 .46 stage in testing new lines. On the other hand. if Soya bean, seeds, flour or products 5. 71 yield is most important. nutritional testing may Sesame, safflower, sunflower 5. 30 be given lower priority. It must be recognized. however. that if the new lines are to be used for OTHER FOODS 6. 25 human food. they must be subjected to critical nutritional tests. including appropriate bioassays. 1963 (National Academy of Sciences-National These tests should establish not only the nutri­ Research Council 1963). tional value of the cultivar but also its freedom The nitrogen content of any plant material is from toxic substances. a mixture of both protein and non-protein nitro­ gen. To arrive at the protein content of a cereal, Total Protein Analysis the nitrogen content as determined is then multi­ Total or crude protein content is normally plied by a conversion factor to give the percentage determined by the Kjeldahl method as specified protein. The conversion factor used is based on by the Association of Official Analytical Chemists the nitrogen which the particular protein contains. ( 1970). This procedure involves the oxidation of The conversion factor employed is not the same the organic matter in the sample. and the reduction for all cereals nor is it always stated in the litera­ of organic nitrogen to ammonium sulphate ture. A recent compilation (World Health Organi­ followed by the quantitative determination of zation 1973) gave the recommended conversion ammonia. For cereals and legume foods the factors listed in Table 5. Tkachuk ( 1969) recom­ digestion mixture of preference is mercuric oxide. mended a conversion factor of N x 5.76 for dipotassium sulphate and sulphuric acid (Associa­ triticale. For samples containing appreciable tion ofOfficial Analytical Chemists 1970). Methods amounts of non-protein nitrogen the protein of protein analysis have been discussed in con­ content may be more accurately estimated by siderable detail by an International Conference in using the summation of the total weight of amino HULSE/LAING: PROTEINS OF TRITICALE 29 acids determined by ion exchange chromatography Because the apparent recovery of protein (sum (see next section). of individual amino acids divided by nitrogen x It may be concluded that the first step in any appropriate factor) varies considerably even evaluation of protein is the determination of among different pure proteins, Knipfel et al. total or crude protein in the sample by the ( 1971) suggested adjusting the total recovery to a Kjeldahl method. For screening samples of limited constant value of90~~- However for protein sources size, semi-automatic micromethods are available. containing a high proportion of non-protein nitrogen much lower recoveries are possible.

Amino Acid Analysis Microbiological A~ays Protein Hydrolysis Microbiological assays are less satisfactory In order to determine the amino acid content than ion exchange chromatography for complete the protein must first be hydrolyzed. There are amino acid analyses. They are, however, con­ no official methods for either hydrolyzing proteins venient for estimating certain individual limiting or for determining their concentration in the amino acids such as lysine, methionine, threonine hydrolysate. Blackbum (1968) has selected what and tryptophan in large numbers of samples. The he considers the most appropriate procedures for authors have obtained satisfactory assays for amino acid analysis. Generally, a weighed, finely the above four amino acids using a commercially ground sample is hydrolyzed in 6N HCI for 20--24 prepared amino acid assay media (Difeo), and a h at 11 O- I I 5°C under nitrogen in sealed glass test 72-h incubation period followed by titration of tubes or in Teflon bottles with screw caps. The lactic acid. Microbiological assays have the hydrolysate is made up to a given volume, advantage that they require relatively inexpensive filtered, and an aliquot is evaporated to dryness equipment which includes an autoclave, an incu­ in a rotary evaporator. The residue is dissolved in bator (or water bath), and general laboratory citrate buffer (pH 2.2) and diluted for subsequent glassware. Although these assays are relatively analysis either by microbiological assay or auto­ simple, the results are affected by many factors mated amino acid analyzer. including age and number of cells in the inoculum, The effects of acid hydrolysis on different variation in temperature during incubation, loca­ amino acids in various plant foods must be tion within the incubator, and contamination with recognized. It is important to obtain full release of other bacterial species. A useful reference book amino acids while keeping their degradation to a is by Barton-Wright ( 1952). Other references re­ minimum (Tkachuk and Irvine 1969). It must be lating to microbiological assays include Schiaffino realized of course that acid hydrolysis yields total ( 1959) and Hom et al. ( 1953). amino acid content and gives no indication of nutritional availability of the amino acid. Since Amino Acid Units tryptophan is completely destroyed during acid The amino acid content of protein is expressed hydrolysis, other methods of hydrolysis must be in various ways. Older tables of amino acid employed when assaying for this amino acid. The content (Block and Weiss 1956) gave amino acid newer procedures use the enzyme Pronase (Cal­ values in grams amino acid per 16 g nitrogen, biochem, Los Angeles, U.S.A.) (Spies 1967; i.e., essentially in terms of percent of protein. Wall et al. 1971). This assumes, incorrectly, a constant conversion factor of 6.25 whereby to convert "nitrogen" to Ion Exchange Chromatography protein (16 x 6.25 = I 00). Other ways include The most satisfactory method for determining micromoles per unit of nitrogen or of protein. amino acid composition is ion exchange chro­ The convention used by the Food and Agriculture matography. Most laboratories conducting amino Organization (World Health Organization 1973) acid analysis use modern automated amino acid is milligrams amino acid content per gram total analyzers employing the ion exchange principles nitrogen or protein. originally developed by Moore and Stein ( 1951 ). Since analyses may not include tryptophan, Available Amino Acids praline, hydroxyproline and cystine, I 00% re­ It may be concluded that hydrolysis of proteins covery of amino acid nitrogen cannot be expected. is probably the most critical analytical step in the 30 MONOGRAPH IDRC-02le

estimation of amino acid content. The method­ a reference standard since it had a known, high ology appropriate will depend on the number of biological value. It was assigned a value of JOO amino acids being investigated and equipment and the assumption was made that the amino acid available. It must also be recognized that in some profile of egg contained the same proportions of foods, particularly those which have been heat essential amino acids as the requirement pattern processed, all the amino acids present may not be of the animal. Since egg apparently has too much fully available. Obviously, there is need for uni­ methionine and cystine (McLaughlan et al. 1959; versal testing and adoption of methods, partic­ Rao et al. 1964) in relation to the ideal amino ularly for protein hydrolysis. Until a universally acid requirement for the growing rat, its use as acceptable methodology is agreed upon some con­ the reference standard exaggerates any deficiency tradictions and confusion must be expected among of these amino acids in proteins under test. Other different determinations of the amino acid con­ problems in the use of chemical score have been tents of cereals and other foods. reviewed recently by Bender (1973). At present a variety of amino acid reference patterns (egg, human milk, and estimated human Chemical Score amino acid requirement pattern) are employed in The quality of protein in a food is dependent arriving at a chemical score. Although human largeiy on the amount of the limiting amino acid amino acid requirements would seem to be the present in the protein. "Limiting amino acid" has logical standard, this pattern is based on exceed­ been defined (National Academy of Sciences­ ingly variable data (Irwin and Hegsted 1971 ). National Research Council 1963) as the essential In 1965 a joint FAO/WHO expert group (World amino acid of a protein which shows the greatest Health Organization I 965a) recommended egg or percentage deficit in comparison with a standard. human milk amino acid patterns as the standards. Various ways of expressing this deficit have been However, a recent joint FAO/WHO Expert suggested. The chemical score method (CS) was Committee (World Health Organization 1973) originally proposed by Mitchell and Block (1946) has proposed a provisional amino acid scoring for predicting the quality of proteins (rom the pattern which contains considerably reduced amino acid profile. Because amino acid require­ amounts of the sulphur-containing amino acids. ments were not established at the time of Mitchell The calculation of CS by this method is shown and Block's proposal, egg protein was chosen as in Table 6 (World Health Organization 1973).

TABLE 6. Calculation of chemical score for cereals, soybean and a mixture of maize and soybean: amino acid content calculated from data in Food and Agriculture Organization ( l 970b).

Maize-soybean World (I : 1 protein Health Wheat Maize Soybean mixture) Organization (1973) Amino Amino Amino Amino Requirement acid acid acid acid pattern content content content content (mg jg (mg/g Chemical (mg/g Chemical (mg/g Chemical (mg/g Chemical Amino acid protein) protein) score protein) score protein) score protein) score

Lysine 55 31 56 27 49 70 127 48 88 Threonine 40 31 77 36 90 42 105 39 97 Methionine and cystine 35 43 123 35 100 28 80 31 88 Leucine 70 72 103 125 180 85 121 105 150 Isoleucine 40 35 88 37 93 50 125 44 109 Valine 50 47 94 48 96 53 106 50 100 Phenylalanine and tyrosine 60 81 135 87 145 89 148 88 147 Tryptophan 10 7 70 14 140 10 100 HULSE/LAING: PROTEINS OF TRITICALE 31

Using the definition given above, the CS for TABLE 7. Available lysine in heated casein-glucose lysine in wheat is (31 -;- 55) x JOO= 56. The mixtures (Rao and Mclaughlan 1967). lowest score indicates both the limiting amino acid and the extent of the amino acid deficiency. Available lysine Lysine is shown to be the limiting amino acid in each of the cereals but threonine is the second Lysine Gross limiting in wheat whereas tryptophan is the second Autoclaving DNFB bioassay Protein time (min) (chemical) (rat) limiting in maize. The information on triticale value (rut) available at the present time is insufficient to 0 6.10 7.09 7.53±0.26 compute a reliable CS along lines comparable to 5 5.62 6.98 6. 22 ±0. 36 those quoted for other cereals. Based upon the IO 3.50 3.71 4.26±0.55 limited analytical data available, triticale would 20 2. 11 1.47±0.58 appear to have a CS for lysine of about 64. It is 40 1.94 0.48±0.29 customary to combine the sulphur-containing amino acids in the one score; these are seen to be •Growth was too erratic to estimate lysine avail­ limiting in soybean protein. It is interesting to bility. note the supplementary relationship when maize and soybean protein are combined. The CS of 88 When a protein source is heated with other for lysine and for methionine plus cystine is better foods, reactions occur between free amino groups than the score of either protein source alone. It in the protein with the sugars, aldehydes and fatty should be pointed out that this pattern places less acids, which may limit the availability of the amino emphasis on sulphur amino acids than do any of acids. The rapidity of the reaction between lysine the previously recommended reference patterns. and glucose during heating is illustrated in Table Despite the variety of amino acid reference 7 (Rao and Mclaughlan 1967). A mixture of patterns which have been suggested, many studies casein and glucose in water was autoclaved for have confirmed the general validity and usefulness varying periods. Available lysine was estimated of CS for expressing protein quality. It is strongly using rats by a direct lysine bioassay and by recommended that the most recent reference Gross Protein Value. The dinitrofluorobenzene pattern in Table 6 should be used. It must be re­ (DNFB) procedure of Carpenter and Ellinger membered however that amino acids in some (1955) for estimating available lysine was also foods, particularly heat processed ones, may not be conducted. Each of the three methods indicated a fully available, in which event the CS which is marked fall in lysine availability with as little as calculated from total amino acids, not total 10 min heating time. Because of its rapidity, the available amino acids, will reflect a higher bio­ DNFB method is valuable in determining avail­ logical value than is in fact the case. able lysine.

Amino Acid Availability DNFB Method There are several causes of reduced amino acid The DNFB method is based on the reaction availability but probably the most common is between DNFB and the free epsilon amino group simply poor digestibility of the food. Nutrients in of lysine in proteins. Unavailable lysine, i.e. when plants may not be completely digested because of linked with sugars or other compounds, is not interference by the cellulosic cell walls. The free to react with DNFB. On acid hydrolysis, the anti-trypsin factor in soybeans is well known product of the reaction between lysine and DNFB, (Liener 1969) but this factor may also be present dinitrophenyl-lysine is released and can be mea­ to some extent in certain cereals (Couch and sured spectrophotometrically. The original method Hooper 1972). Moderate heating inactivates the is not satisfactory, however, for high carbohydrate trypsin inhibitor, thereby increasing the digesti­ foods such as cereals (Conkerton and Frampton bility of the protein. Excessive heat treatment, 1959) and modifications have been proposed. Rao however, renders the protein resistant to digestive et al. (1963) separated DNP-lysine quantitatively enzymes, and even in roller dried milks there is from interfering components by ion exchange frequently a reduction in the availability of lysine chromatography; this modification has been used (Mauron 1961). successfully on plant products (Wall et al. 1971). 32 MONOGRAPH IDRC-021e

Enzymatic Hydrolysis processed foods containing sugar than in cereal Since amino acid availability may be related to products that are subjected to relatively mild reduced susceptibility to enzymatic activity, several heating. Nevertheless, amino acid availability is methods, based on the hydrolysis of proteins by one of the chief criticisms of CS which assumes enzymes have been suggested, e.g. Mauron ( 1961 ). complete availability. Thus, the determination of The basis of these methods is to simulate the available amino acid content of protein is an sequence of proteolytic enzymes that operate in essential test for the analysis of processed foods. vivo. The sequence followed is usually digestion Since factors such as availability may influence with pepsin at pH 2.0 followed by trypsin and the apparent value of protein in foods, it is im­ erepsin at pH 8.0. Hydrolysis of protein is usually portant that there be a critical evaluation of not complete even though it is continued for 2-3 protein by rat bioassays. Several methods have days. A variety of methods (Mauron 1961) have been suggested and their advantages and dis­ been employed to assess the extent of amino acid advantages critically discussed (National Academy release, but none of these methods has achieved of Sciences-National Research Council 1963: widespread acceptance. Enzymatic digestion com­ McLaughlan and Campbell 1969; Pellett 1973). bined with a microbiological assay for a specific amino acid such as methionine was recommended by Boyne et al. (1967) to assess amino acid avail­ Biological Quality ability in high protein supplements for the animal feed industry. Protein Efficiency Ratio One of the most widely accepted procedures for Microbiological Methods determining protein quality is the Protein Effi­ Ford (1962) reported that the proteolytic ciency Ratio (PER) method in which rats are fed bacterium Streptococcus zymogenes NCDO 592 under specified controlled conditions for a pre­ was suitable for estimating the availability of scribed time. The PER is defined as the ratio of methionine, leucine, isoleucine, valine, tryptophan, body weight gain (in grams) to the weight (grams) arginine and histidine. The bioassay system mea­ of protein consumed. Since the results obtained sured the amount of each specific amino acid are influenced by factors such as the age of rat, released enzymatically by a preliminary treatment length of assay period, level of protein and sex of of sample with papain and by th·e exocellular rat, these conditions must be standardized. Such enzymes secreted by the test organism. The method standardized procedures are set out in official was subjected to a collaborative assay and it methods in the United States of America (Associa­ appeared suitable for measuring amino acid tion of Official Analytical Chemists 1970) and availability (Boyne et al. 1967). Unfortunately the Canada (Food and Drug Laboratories 1966). In assay organism does not require lysine and is both these assay procedures, the results are com­ therefore unsuitable for estimating available puted in comparison with a standardized casein lysine; Ford (1964) recommended Streptococcus sample to which is assigned a PER value of 2.5. durans for this purpose. These methods are 6asi­ The standardized procedure prescribes the age of cally sound but some difference of views exist the male rats to be used, a 4-week assay period regarding the preliminary enzymatic treatment on an ad libitum diet containing 9 or 10% protein (Szmelcman and Guggenheim 1967). on a dry weight basis and adequate in other Shorrock and Ford (1973) reported experience essential nutrients. with the protozoon Tetrahymena pyriformis for In view of the influence of the factors noted the assay ofavailable lysine and methionine. Using above, the term PER should be applied only to the method of Stott and Smith (1966) they con­ results obtained from standardized assays. In cluded that while the Tetrahymena assay can reporting data, any variations from the official predict accurately the results of growth tests with methods should be clearly recorded. If animals rats, it would be premature to recommend its other than rats as prescribed are used for protein general use to replace the bioassay. evaluation, the term PER should not be used. Although the PER method has achieved wide­ Effect of Heat Processing spread acceptance, it may be criticized from several It may be concluded that the availability of aspects. Thus, it has been pointed out that (a) amino acids is much more of a problem in heat- there is not a proportional relationship between HULSE/LAING: PROTEINS OF TRITICALE 33

80 usually does for casein. It is clear that the highest ,/______J 70 dietary protein level gives the largest PER provided the point falls on the linear portion of 60 the growth response curve. Over this portion of the curve the protein required for body tissue maintenance is of lesser importance than that ~ 40 z required for growth. "r u 30 ./'""'""" Net Protein Retention To make allowance for body tissue replace­ 10 ment and maintenance, the Net Protein Ratio (NPR) assay was proposed by Bender and Doell 0 ----~;/-----·~ (1957). The NPR is equal to the mean weight -10 / / gain of the test group plus the mean weight loss of a group of rats fed a non-protein diet divided by 2 4 6 8 10 12 14 16 18 20 PROTEIN CONSUMED (9) the average protein consumed by the test group. Thus, for lactalbumin (Fig. I) fed at a 9°1,, protein Fig. I. Weight change of rats fed three /erels of -o- /actalbumin and v•heat gluten. level the NPR = (75 + 20) 20 = 4.74. For wheat gluten the NPR at the 9% protein level is 2.21 and one PER value and another, i.e. a PER of 2.0 is at 7.5~/,, protein the value is 3.54. Thus NPR is not twice as good as a PER value of 1.0, (b) the also influenced to some extent by the percentage results may be influenced by level of food intake, protein level. Details of the procedure are given (c) maximum PER values are not obtained at the in Recommended Methods at the end of this same dietary protein level for different proteins, chapter. (d) the method makes no allowance for body tissue maintenance but assumes that all protein con­ Net Protein Utilization sumed is used for growth. In spite of these short­ The Net Protein Utilization (NPU) assay comings, the PER method, until recently, has described by Miller and Bender (1955) is similar been considered the simplest and most generally to NPR except that body nitrogen rather than applicable method (McLaughlan and Campbell body weight is the parameter measured. Bender 1969). Its apparent simplicity may, however, and Doell ( 1957) found that NPU and NPR have encouraged persons lacking a full apprecia­ methods correlated closely (r = 0.99) among a tion of its inherent complexities to report results variety of 35 foods. Because of the carcass obtained without due regard to the standardiza­ nitrogen analyses required, the NPU assay is not tion necessary. Because of its widespread accep­ as amenable as the NPR assay to routine testing tance a description of the method recommended ofa large number of samples and there is evidence for use in Canada is given in Recommended that it may be more variable (Chapman et al. Methods at the end of this Chapter. 1959). Some of the factors which influence PER assays are shown in Fig. I in which body weight change is plotted against protein consumed at three levels Slope Ratio Assay for lactalbumin and wheat gluten. Since PER is Hegsted and Chang (I 965a) proposed the slope defined as the ratio of grams of weight gain ratio (SR) assay in which determinations are made d.ivided by grams of protein consumed, the PER at several dietary protein levels. The slope of the at 9% protein level for lactalbumin is 75.0 -o- 20.0 = growth response line (i.e. the straight portion from 3.75. PER values at 3, 5 and 7% protein levels zero protein intake up to the point where the line were 0.25, 2.95 and 3.93% respectively. PER begins to curve towards the horizontal) of the test values for wheat gluten at 7.5, 9 and 15% protein protein is expressed as a percent of the correspond­ levels were 0, 0.41 and 0.60, respectively. Although ing slope for a lactalbumin reference protein. This PER is arbitrarily measured at the 9-10% level method has the advantage that the assay defines of protein, this level does not give the maximum the slope of the response curve and furnishes an value for either lactalbumin or wheat gluten but estimate of the error of the assay and its validity. 34 MONOGRAPH IDRC-021e

In Fig. I the slope of the line up to the 7° 0 threonine in peanut protein. Thus, the results of protein (lactalbumin) intake is (59 + 20)-'- 14.5 = bioassays depend partly on the level of protein in 5.45. Evidently, the slope does not change with the diet as well as upon which amino acid is different protein intakes up to the point at which limiting in the protein. Particular care needs to be the growth response curve begins to flatten. The taken therefore in assessing quality of cereal pro­ growth response curve for lactalbumin could be teins. Despite the shortcomings of rat bioassays, extended to cut the Y axis at - 20 g (i.e. the weight results on a series of protein sources including loss of the group of rats fed the non-protein diet). poor, moderate and good proteins showed close The growth response curve for wheat gluten correlation between results of bioassays using cannot be extended to cut the Y axis at - 20 g rats and humans (Campbell and McLaughlan and consequently the SR assay for wheat gluten 1970; Bressani et al. 1973). is invalid. Similar difficulties have been experienced with other lysine-limiting cereal proteins (Said Bioassays-Conclusions and Hegsted 1969; Yanez and McLaughlan It is concluded that in spite of the inherent 1970). This problem seriously limits the usefulness difficulties of rat bioassays, they must be con­ of the SR assay. sidered an essential part of any comprehensive It was found from an analysis of available data testing program. While the PER method is simple (McLaughlan and Campbell 1972) that the slope and reproducible, there appear to be significant ratio assay could be improved by omitting the advantages to the use of the GS assay in terms of group of rats fed the non-protein diet. This finding validity and reliability. Protein values remain was also made independently by Hegsted ( 1972) constant at different levels of protein intake. from the results of a collaborative assay. This Although further investigation is needed, the GS modified slope ratio assay which we have called assay is recommended at this time as the most the "'growth slope" (GS) assay is essentially the reliable method for the assay of the protein quality same as the Nitrogen Growth Index of Allison of cereal grains and legumes. Where the quantity (1964). of cereal or legume to be assayed is small, a single In the GS assay the true slope of the response level NPR assay using five animals in ·a IO-day line is used. The slopes (Fig. I) for lactalbumin assay, is a valuable preliminary screening test. and wheat gluten would be 5.45 and 0.86 respec­ tively giving a GS assay value for wheat gluten of Whatever method is selected, it must be standard­ ized and carried out according to recognized 0.86-'- 5.45 x 100 = 16. There is at least one procedures. Details of GS assay, NPR and PER problem with the GS method; the response curve methods as recommended by the authors are for peanut protein tends to have the same slope, given in the Recommended Methods at the end of and thus the same GS value, as casein yet it is this Chapter. Variations in these procedures inferior to casein for maintenance purposes. This should not be used unless carefully tested and peculiarity is believed to result from a relatively specified. high requirement of the rat for threonine (and possibly for sulphur-containing amino acids) for maintenance purposes. This aspect of the GS method requires further study. In view of the need to standardize and recognize protein method­ Rapid Screening Methods ology officially, it is urged that this method be so evaluated at an early date. Microbiological Methods It is also evident from Fig. I that wheat gluten Because of the need to test large numbers of appears to be equivalent to lactalbumin at low small samples the plant breeder has a particular protein intakes. The reason is that the relative interest in rapid screening methods. From time lysine requirement for maintenance is low com­ to time various methods utilizing the growth of pared to its requirement for growth of new tissue. protozoa, bacteria or insects, primarily because Consequently some proteins, particularly cereal of their apparent simplicity, have been suggested protein, may be more valuable for body main­ as being useful for distinguishing between various tenance purposes than for growth (Campbell and levels of protein quality for specific foods. Since McLaughlan 1970). The opposite may be true for the protozoon Tetrahymena pyriformis requires HULSE/LAING: PROTEINS OF TRJTICALE 35 the ten amino acids essential for the rat and is protein molecule. Consequently a highly positive capable of digesting intact proteins, its use seemed correlation exists between the amount of azo dye to offer particular possibilities. The collaborative bound and the number of basic groups associated assay reported by Boyne et al. ( 1967) made it with the protein in the test sample. Dye-binding clear. however. that this organism requires further methods have been used for several purposes study before it can be recommended as a rapid including the determination of total protein screening assay for protein quality. Haenel and content of foods (Cole 1969) and have also been Kharatyan ( 1973) have stated that results from used to detect changes in availability of lysine. Tetrahymena and from other microbiological Since the methods are rapid and inexpensive methods for determining relative nutritive and may be automated, they have found favour in value should not be considered an absolute testing new lines of crops. They have been measure of protein quality without additional officially recognized for the analysis of protein confirmation. in milk and milk products (Association of Official Halevy and Grossowicz ( 1953) and Teeri et al. Analytical Chemists 1970). The UDY dye method ( 1956) among others, have used S. ftu'calis for for crude protein in wheat and flour has been similar purposes. The problems inherent with such approved by the American Association of Cereal assays have been clearly pointed out by Rogers Chemists ( 1968). et al. ( 1959) who indicated that these methods were Lakin ( l 973a) has recently reviewed the subject valid only when the standard and sample were in detail and cited examples of the use of dye­ limiting in the same amino acid. Since this is often binding for the selection of high lysine strains of not known or needs evaluation, the use of the S. cereals and legumes (Munck et al. 1969, Kaul faecalis method for screening purposes is not et al. 1970) and for studying the effects of factors recommended. affecting harvesting and storage of grain (Mossberg Shorrock and Ford ( 1973) investigated the use 1970). Lakin ( l 973b) has also indicated that of both Tetralnmma pyriformis and S. Znnogenes although dye-binding procedures may be used as and reported that while both could be used for routine methods to determine protein content of determining available methionine, the :::rmogenes a wide range of foods, they are empirical pro­ assay seemed simpler. They also concluded that cedures and a reference method must be used for Tetrahrmena was useful for determining available calibration purposes. It has been found that the lysine under controlled conditions. It is suggested calibration will hold as long as the standardiza­ that neither bacteria nor protozoa are useful as tion of the dye-binding technique is maintained non-specific screening methods. They can be and the character of the samples remains much more effectively directed toward the esti­ unchanged. mation of individual amino acid content or Lakin (1973b) considered CI Acid Orange IO available amino acid content using appropriate to be the best dye for the direct evaluation of media. protein quality and it may be used, in a modified procedure. for the determination of available lysine. Dye-Binding Methods It must be emphasized that dye-binding is an Dye-binding procedures have been used for empirical method and since the results are pro­ some time for the estimation of protein and the portional to the basic amino acids (e.g. lysine, evaluation of its nutritional quality. Fraenkel­ histidine, arginine) present. variations in the Conrat and Cooper ( 1944) described how dyestuffs percentages of these basic acids among samples can be used to determine the number of basic of a given cereal may cause discrepancies when groups present in proteins. The precise nature of dye-binding is used to determine total protein. the dye-protein interaction is obscure but the Furthermore, one cannot use dye-binding pro­ dominant mechanism is believed to be an electro­ cedure to determine total protein and again to valent association between the dye anions and determine lysine as percent of protein in the same electro-positively charged sites in the proteins. sample. Obviously, much more work needs to be These charges occur in the terminal amino groups done on these methods before they can be used and in the basic groups associated with the histi­ reliably to predict the protein value of new culti­ dine, arginine and lysine residues present in the vars. 36 MONOGRAPH IDRC-02le

Biological Methods greater uniformity than those derived from voles, The need for a rapid, simple biological test has a considerable amount of work needs to be done been widely recognized. It has been suggested by before the mouse assay can be considered ade­ several workers (Elliot 1963; Shenk et al. 1970) quately standardized and reliable. that the meadow vole (Microtus pennsylranicus) Since rye may contain certain toxic or un­ has unique advantages for the biological evalua­ desirable substances, e.g. ergot and resorcinols, tion of certain plant materials. The vole has also the possibility exists that such substances may be been suggested as a means of screening new lines present in triticale (see Chapter 4). The vole of triticale in that (I) very small samples can be has been suggested as being useful in their detec­ tested quickly, and (2) the vole gives simultaneously tion. It should be emphasized, however, that there an estimate of protein value and an indication of is no single level animal test that has any meaning the presence of toxic factors. in this regard. One approach that has been found In fact, the vole assay suffers from several very useful (Campbell 1963) is to feed protein supple­ serious defects (B. E. McDonald and E. N. Larter ments or concentrates at various levels up to 1972 unpublished data). Data provided by 15-18% protein in a rat growth assay. If there is McDonald and Larter and cited in Table 8, show evident growth retardation at the high levels, it that the range of variation within groups of voles may be suspected that the food or protein has is very wide, rendering the vole ineffective as a some toxic or undesirable properties. Unfortu­ test animal. It is also evident that voles cannot nately, this method is not too useful for cereals differentiate between the quality of protein in which contain relatively low levels of protein. foods as divergent as casein and cereals. Little is Other suggestions are given by the Protein Advi­ published about the physiology of the vole and sory Group (I 972b). practically nothing appears to be known of its amino acid requirements. Thus, the meaning of Rapid Screening-Conclusions the vole assay as described is in considerable In general, it may be concluded that there is no doubt in terms of human needs. There are no relatively simple, reliable, screening method for data to describe the reaction of the vole to any evaluating protein quality. There is an urgent "toxic" substance which may be present in new need for such a method and research must be lines of triticale. The vole does not appear to directed at once towards this matter if programs thrive under normal animal laboratory conditions for the development of new plant cultivars are but requires a very moist atmosphere which is to be effective. Non-specific, non-standardized difficult to maintain. In view of these limitations. assays of any type whether biological, chemical or it seems clear that the meadow vole cannot be microbiological, no matter how simple and easy recommended for the biological testing of new to apply, should not be used until their meaning, lines of triticale or other cereals intended for usefulness and limitations have been clearly human use. established, and the techniques for using them The laboratory mouse was also investigated as carefully standardized. a test animal by McDonald and Larter (un­ published data) and while the results exhibited Bioassays with Humans The classical method of determining biological TABLE 8. Results of growth assays using voles on four value as a measure of protein quality was that of samples (McDonald and Larter 1972 unpublished). Thomas (1909) and Mitchell (1923-24). However, because this method requires measurements of Vole nitrogen intake and both faecal nitrogen and preliminary urinary nitrogen while on both a test diet and a Index non-protein diet, it is not suitable as a routine test Sample (7-day assay) SD for assessing protein quality. When carried out with human subjects under closely controlled con­ Casein 0.69 0.92 ditions, however, this method still remains the Wheat 1.16 0.79 Triticale 1.07 1.09 ultimate standard for evaluating protein quality. Rye 1.62 0.69 Another approach is based on the growth and development of young children consuming the HULSE/LAING: PROTEINS OF TRITICALE 37 new food as almost the sole source of protein sideration must be given to the impact that an (Graham et al. 1966). Unfortunately, the growth effective program may have on the overall test can seldom be used because of its long nutrition of the particular area or country. Since duration. the prime purpose of any program aimed at foods The most widely used method is nitrogen balance for human use is to improve the quality and in which the nitrogen intake and nitrogen losses in quantity of the foods, in this case the quality and urine and faeces are measured. The procedure is quantity of protein, it is most important that there particularly useful when conducted with young, be close collaboration between plant breeders, healthy children. Scrimshaw et al. ( 1958), Bressani nutritionists and analysts to ensure that the pro­ and Viteri (1970) and Bressani et al. (1973) have gram is nutritionally sound and that appropriate used nitrogen balance to measure the nutritive methodology for evaluation is employed. The value of various proteins. The technique is partic­ following scheme is suggested for the evaluation ularly valuable when applied as the nitrogen bal­ of protein in new cereal lines. ance index in which the slope of the response line is Procedure I -For very small samples: determined. The latter workers concluded that this (I) Determine total protein by Kjeldahl method approach is particularly useful to evaluate protein using Association of Official Analytical quality with human subjects, and that comparative Chemists (1970) procedure. tests with rats appear to correlate well with human (2) Determine amino acid content using ion ex­ data. Kies et al. (1965) have applied the method change chromatography (Blackburn 1968), widely in studies with cereals. Graham et al. or autoanalyzer. ( 1966) have used changes in plasma albumin in (3) Determine limiting amino acid content conjunction with either growth or nitrogen balance using FAO/WHO reference pattern (World in assessing protein quality in infants. Health Organization 1973). Although many studies, e.g. Young and (4) If sufficient sample is available conduct an Scrimshaw (1972) have explored the possibility abbreviated rat assay, e.g. five animals per of using changes in levels of free amino acids in group in IO-day NPR assay (see Recom­ blood as a basis for estimating protein quality, mended Methods at the end of this chapter). no practical method has evolved. The level of lysine in the blood may indicate whether or not Procedure 2-For adequate sample size: lysine is the limiting amino acid in a cereal diet Use Multilevel rat assay, i.e. Growth Slope but it gives no quantitative measure of the extent Assay (see Recommended Methods at the of the deficiency. Blood urea levels have also been end of this chapter). suggested as a measure of protein quality but as Obviously any food for human use must not Eggum (1973) has pointed out, the experimental only have appropriate nutritional value, it must conditions need much further standardization. also be free of toxic or harmful substances It is concluded that the nitrogen balance index (Protein Advisory Group I 972b). No test for method for the testing of proteins with humans is protein value will give an evaluation of the cumbersome to use routinely. The main value of presence of such substances. If digestible protein the procedure appears to be in the validation of is present in sufficient concentration, a multilevel rat bioassays and the final assessment of new rat growth test should demonstrate the presence cultivars. A more suitable method for human of toxic factors. evaluation of protein quality is urgently needed and particular attention should be paid to the analysis of body fluids. Summary I Any plant improvement program must be Suggested Testing Procedures designed to meet the nutritional needs of the In any program for the improvement of the particular country or area for which the crop is nutritional properties of new lines of cereals and being designed. legumes, it is essential that the objective of the 2 The first step in the evaluation of protein is program be clearly established and understood. the determination of crude or total protein by In determining the need for new cultivars con- approved methods. 38 MONOGRAPH IDRC-02Ie

3 The second step is the estimation of amino Recommended Methods acids, particularly the limiting ones, preferably PER Assay by ion exchange chromatography. The use of Chemical Score is helpful in rating proteins, and Animals Use male weanling rats of a single determining limiting amino acids. strain, 21-23 days ofage. Place on a commercial rat chow diet for 2 days. Use groups of IO rats 4 Tests of availability are necessary to estimate for each diet with a weight range not greater the actual amount of amino acids that may be than 15 g. Distribute rats to various groups so utilized by the human. Availability may be that mean starting weights of all groups are reduced, particularly in proteins processed by similar. heat in the presence of sugars. Diets Use a basal diet of the following com- Biological testing is an indispensable part of 5 position on an air-dried basis: maize starch 81, any protein evaluation program. Where test maize oil IO, non-nutritive cellulose 5, salts samples are adequate the Growth Slope Assay (Bernhart and Tomarelli, 1966) 3, vitamin appears to be the best available procedure. mixture (Beare-Rogers and Nera 1972) I. (Details of the assay are described in Recom­ Incorporate the test protein in the diet at the mended Methods.) expense of corn starch to I0°fo protein (nitrogen 6 There is no simple, specific, reliable screening x 6.25 or other appropriate factor). The pro­ method for evaluating protein quality. Non­ tein content of the final diet should be within specific chemical, microbiological or biological the range 9.7- I0.3% determined by Kjeldahl screening methods are not recommended unless analysis. they have been very carefully standardized and Assay period 4 weeks. their significance has been established and under­ stood. Cages Use individual screen bottom cages provided with feeders which will reduce food 7 There is no simple screening method for esti­ spillage to a minimum. mating toxic substances in foods but where applicable multilevel rat growth tests are useful. Parameters measured Weigh animals weekly at the same time each day. Estimate food con­ 8 There is urgent need for the development and sumption, taking into account wasted food. standardization of rapid, preferably non-destruc­ tive, tests applicable to small samples of grain Casein control Include a reference standard which will reliably predict protein value. group of rats given casein (ANRC high nitrogen casein, Sheffield Chemical Co., Norwich, N.Y.) 9 Because of the difficulties inherent in evaluat­ in place of the test protein. ing protein in foods, there is need for close col­ laboration among plant scientists, nutritionists Calculation PER= Weight gain (grams)/Pro- and analytical chemists. tein consumed (grams). Assume that casein has a PER of 2.5 and correct the PER of the test 10 In view of the multiplicity of methods avail­ food to a value of 2.5 for casein. able for chemical, microbiological and biological testing of protein value, there is an urgent need for the comparative standardization, selection and NPR Assay official recognition of demonstrably reliable Use animals, diets, and cages as for the PER methods. assay. In addition to the test groups include a The need for standardization of methodology control group of rats fed the unmodified ration for evaluation of plant proteins for human use is (i.e. the non-protein diet). If the sample size is a subject the authors would refer as a matter of limited, use only 5-6 rats per test protein. The urgency to the Protein Advisory Group of the initial weights of rats in the two groups should be United Nations and to the International Unions of matched to within I g. After I 0 days on the diets Pure and Applied Chemistry, Nutrition Sciences, calculate for each pair of rats, the NPR for the and Food Science and Technology. test protein. HULSE/LAING: PROTEINS OF TRITICALE 39

NPR = (WG + WL)/P Assay period 2 weeks. where: WG =weight gain of test animal (g) Parameters measured Weight changes and WL =weight loss of control animal (g) food consumption (as for PER). P = protein consumed by test animal (g ). Calculation of GS Assay Prepare a table con- GS Assay taining weight changes, food intake and cal­ Animals As specified for PER. culated protein (or nitrogen) intake for each animal. Plot mean group data for weight Diets As specified for PER except for protein change against protein or nitrogen consumed content of diets. Test each protein at 3 levels for both standard and test protein. Calculate added at the expense of corn starch. slope from linear portion of each line, and Reference Protein Use lactalbumin at protein express the slope of the test protein as a per­ levels of 2.5, 5.0 and 7.5%. Test higher quality cent of the slope of the reference standard proteins at 3, 6 and 9%. Test proteins of lesser (lactalbumin). Appropriate computer programs quality at 4, 8 and 12, or 5, IO and 15% of the may be used for calculating results, testing for diet to obtain linear dose-response relation. linearity and giving fiducial limits of the assay. n1e p/0111 breedl!r's trl!atment oj the .fl'edlmg jmm tilt' first cross "Ith colchicint hos op1med the door to thl' derelopment of improred lines Chapter 3

GENETIC. VARI ET AL. ENVIRONMENT AL ANO AGRONOMIC FACTORS

Introduction foods and biological data on proteins. and using more selective criteria than Coons. listed 65 This chapter contains a review of the literature references to wheat and wheat products, ten in which (a) genetic and hereditary factors, (h) references to rye, and one reference to triticale. varietal differences and (c) environmental in­ The International Wheat and Maize Improve­ fluences and agronomic practices are related to ment Centre (CIMMYT 1971) has published a the nutritional value of the proteins of triticale three-volume bibliography of wheat. Deodikar and its parents wheat and rye. Every plant is the ( 1963) prepared a review on rye which included result of complex interactions among these factors over 1,000 references and in his preface ac­ and possibly other. as yet unidentified, influences. knowledges his considerable reliance on the publi­ Consequently, though a serious effort has been cation Plant Breeding Abstracts. Pomeranz ( 1971 a) made to separate each of the above defined also includes data on the nutritional value of influences in the text which follows, a clear-cut wheat protein and Aykroyd and Doughty ( 1970) separation and distinction among them is not reviewed the role of wheat in human nutrition on easily achieved and. therefore. a considerable behalf of the Food and Agriculture Organization degree of overlap will be evident. The literature of the United Nations. reviewed is presented under the headings of the The cultivation of triticale has been discussed cereals with which the text is concerned and by Briggle ( 1969) and Zillinsky and Borlaug the effect of the above influences, (a). (b) and (c), (197la. b). The cultivation of wheat has been upon the total protein content. the amino acid reviewed and discussed by Schlehuber and Tucker composition, and the biological value of these ( 1967) and the cultivation of rye by Shands ( 1969). cereals. Moran ( 1959) reviewing the nutritional signifi­ The nutritional quality of any cereal protein is cance of the then recent work on wheat. flour and dependent upon its biochemical composition bread. stated that the differences in the composi­ which in turn is influenced by its genetic back­ tion of wheat were determined largely by the ground. the environment in which it is grown, the environment in which the wheat was grown and agronomic practices by which it is cultivated and only to a small extent by heredity. The main its freedom from infection and infestation. object of the breeding and intercrossing of wheat No earlier reviews covering the literature in varieties was not to improve the biochemical precisely this form have been found. Coons ( 1968) composition; when such improvement occurred. published a bibliography of selected references on it was only incidental to improvement in yield cereal grains in protein nutrition, listing over 500 capability and resistance to disease. references to papers on wheat and over 50 references to papers on rye, available in English. and published between 1910 and 1966. No Genetic and Varietal Factors references to triticale were included. The Food and Plant breeding consists essentially of (a) the Agriculture Organization ( l 970b ). in its com­ establishment of plant populations within which pilation of data on the amino acid content of the probability of desired character combinations

41 42 MONOGRAPH IDRC-021e

occurring is high and (b) the identification and cereal grains to provide more than 80% of their selection from within these populations of those dietary calorie and protein sources, efforts to parent plants possessed of the characteristics discover varieties of higher biological value with­ desired. While it may not be excessively difficult out detriment to yield, disease resistance and to assemble in a single genotype those character­ adaptability, must be intensified. istics which are inherited in a simple manner, a problem of considerable complexity arises when HIGH LYSINE MAIZE the objective is to combine several characteristics The discovery of high lysine mutants in maize each of which is controlled by a number of genetic gives hope that the biological value of the proteins factors. Such genetically complex characteristics of triticale, wheat and other cereals may prove include yield, resistance to pests and disease, susceptible to improvement by genetic manipula­ environmental adaptability, and biochemical com­ tion. During the early I 960's the amino acid position. To bring about the simultaneous combi­ analysis of a large number of maize varieties led nation of as few as 20 desirable genetic factors to the discovery of two important mutants, within a single genotype by conventional inter­ opaque-2 (Mertz et al. 1964) and ftoury-2 (Nelson crossing would require the planting of a popula­ et al. 1965). Both mutants contained substantially tion covering an immense area of land and of a larger proportions oflysine and tryptophan which, size impossible to screen. Consequently, in select­ for maize, are the limiting amino acids. Subse­ ing the parent population for a prospective new quently maize breeders have successfully trans­ variety, the individual entries chosen usually ferred both opaque-2 and ftoury-2 genes into differ in relatively few genetic factors. Thus the "high lysine" hybrid lines and into varietal deliberate combination of many predetermined populations. This discovery has excited wide­ genetic factors within a single plant requires a spread interest among plant scientists in the very prolonged breeding process. possibility of improving the amino acid balance Even using the techniques of population breed­ in other cereals by genetic manipulation. The ing, in which composites of freely intercrossing opaque-2 and ftoury-2 mutant genes appear to individuals are planted and the best of their suppress partially the synthesis of the nutritionally progeny selected to form the next composite, a unsatisfactory alcohol-soluble prolamin fraction simultaneous combination of complex character­ and to stimulate the synthesis of nutritionally istics is difficult, each major characteristic having more important proteins in the water-soluble to be selected from one or more composite albumin and salt-soluble globulin fractions. As populations. would be expected, the compositional differences Largely for economic reasons, the plant occur in the storage proteins of the endosperm; breeder's primary concern, traditionally, and no difference is apparent between the germ protein until comparatively recently almost exclusively, compositions of the traditional varieties and the has been to select high yielding, healthy, disease high lysine mutants. Since in most known varieties resistant and environmentally adaptable cereal the pro Jamin fraction represents about 45% of the grains. Some attention has also been given to the protein of wheat (Triticum vulgare) and about 40% technological properties demanded by processors of the protein of rye (Secale cereale), a genetic and consumers but among many cereal consuming mechanism, similar to that in maize, by which to communities processing technologies have de­ improve the nutritional quality of wheat and rye veloped which best-Suit the properties of the cereal proteins would represent an important discovery grains locally available. To demand of the breeder (Nelson 1969). Single identifiable genes with a in addition to other essential characteristics, ~ "high-lysine" effect similar to the opaque-2 and significant change in the biochemistry, particularly ftoury-2 genes in maize have not, at the time of in the amino acid composition, of a cereal grain, writing, been identified in either wheat or rye inevitably adds to the complexity and cost of the (Mertz 1971; Johnson and Mattern 1972). breeding program and may, at least for the short or intermediate term, require a compromise with other established characteristics. Nevertheless, GENETIC CONTROL OF COMPOSITION particularly for the benefit of people in the The improvement of plant protein quality by developing countries, many of whom rely upon genetic manipulation produces an intrinsic and HULSE/LAING: PROTEINS OF TRITICALE 43 integral improvement in that it raises the plant"s cereals are rich in the amino acids praline and potential to produce protein of higher quality in glutamine but comparatively poor in lysine. It is response to favourable environment and agro­ the prolamin fraction which largely contributes nomic management. There seems little doubt that to wheat its unique gluten-forming and tradi­ if its protein were qualitatively and quantitatively tionally desirable bread-making properties. satisfactory. a cereal diet could provide for most Technologies developed recently permit bread human needs. to be made from flours much poorer in gluten­ The fundamental aspects of genetic control of forming proteins and richer in .. high lysine"" protein composition in cereal grains has been proteins than was formerly considered possible elegantly discussed by Nelson ( 1969). The compo­ (Axford et al. 1963 ). Therefore. the apparent con­ sition of plant protein depends upon a genetic flict between technological properties as con­ information system consisting ofa unique sequence tributed by high prolamin components. and oftrinucleotide codons (DNA) which is translated nutritional value from higher lysine components. through a complementary sequence of trinucleo­ may no longer be of serious consequence. tide codons of .. messenger"" RNA into the pre­ determined amino acid sequence. Each codon PROLAMIN SYNTHESIS specifies a particular sequential amino acid. the Since the synthesis of prolamin in maize endo­ amino acid sequence in any given protein being sperm can be genetically suppressed there is hope invariate. Each protein is believed to be synthe­ that comparable mutant genes may be found sized amino acid by amino acid. the process whereby to suppress prolamin synthesis in wheat. starting from the N-terminal and finishing at the rye and other cereal endosperm proteins. The C-terminal end. Since even a single change in presence of a semi-dominant mutation such as amino acid sequence can prove lethal or at least occurs in the floury-2 maize in one of the wheat or injurious to the organism. the established amino rye genomes could cause a significant depression acid sequence tends strongly to be conserved in of prolamin synthesis and should be identifiable. healthy plants. This is particularly true of the As already mentioned. no such mutation has been essential proteins present in the embryo and those reported. It is possible that a search for such which control the photosynthetic and other vital mutations among rye varieties might well prove processes in the leaves. Consequently any hope of more fruitful. improving the amino acid pattern of a plant seed It is recorded by Favret et al. ( 1970) that when protein must be focussed upon the storage prolamins are specific they are regulated by protein of the endosperm. the composition of Mendelian genes which act directly in the endo­ which is less critical to the plant"s survival. sperm tissue and are not influenced by the mother In every organism each cellular component is genotype. in sharp contrast to the potential for provided with its own unique spectrum of proteins. total protein production in the grain which is While the amino acid sequence of each protein strongly influenced by the mother genotype. remains constant. the proportion of the different As already stated. prolamin is comparatively proteins within the spectrum may vary. By what rich in praline whereas lysine is one of the group control mechanism the ratio in which the various of basic amino acids which are present in low proteins are laid down is by no means clearly proportion in prolamin. The ratio of basic amino understood. acids to praline in the endosperm varies among species and with the age of the plant. the ratio CEREAL PROTEIN CLASSIFICATION falling as seed endosperm fills out since prolamin Osborne ( 1924) provided the classification of production takes place largely after the other seed proteins which. to all intents and purposes. storage proteins have been laid down in the still holds good today. Seed proteins contain endosperm. albumins (water-soluble). globulins (soluble in In selecting for wheat and rye genotypes of saline solution). prolamins (soluble in alcohol). improved biological value. it might be useful to and glutelins (soluble in dilute alkali). Only determine the basic amino acid: pro line ratio. among the seeds of the Gramineae. which include Geneticists might also explore the activity of wheat and rye. are prolamins and glutelins found prolamin-regulating genes by examining the in any quantity. The prolamin fractions of all electrophoretic patterns for the appearance and 44 MONOGRAPH IDRC-02le disappearance of specific desirable and undesirable which it is grown is an important influence upon proteins. whether or not the plant achieves its full genetic potential for protein production. There exists an IMPROVEMENT IN SEED PROTEIN immense literature which relates to the influence of In selecting for high protein genotypes it would agronomic variables upon yield, composition and appear to be more meaningful to select on the other characteristics of wheat and, to a lesser basis of protein content per seed rather than pro­ extent, of rye. Since much of the data recorded is tein as percent dry matter. The latter is strongly specific for the location, or closely similar loca­ influenced by seed weight and kernel characteristics tions, in which the experiment was carried out, which in turn are known to be influenced by no attempt has been made to review such literature maturity, environment and agronomic practices. comprehensively in this publication. Other means by which the protein quality of a In addition to the influence of environment, cereal grain might be improved would be to in­ fertilizer and irrigation regimes. and the control crease the size of the embryo relative to the size of of pests and diseases, other factors, including the the endosperm, or to increase the depth of date of planting, can materially influence protein aleurone layer (Nelson 1969). Both the embryo content. It has been reported, for example, that and its surrounding scutellum, and the aleurone triticale lines planted during the spring contained layer contain proportionally more protein nitro­ significantly higher protein contents than the same gen than the endosperm and, in both instances. lines planted in the autumn. the protein is of superior biological composition. Exceptional progress has been made during the In many countries, except for a few specialty past year in the CIMMYT-Manitoba project in breads, the embryo is normally excluded from improving the fertility, the seed density, kernel flours used for bread and alimentary pastes. characteristics and the overall yield capability of Modern roller and abrasive milling processes can triticale. At CIMMYT triticale yields in excess of however be adjusted to include in the flour sub­ 8300 kg/ha (var. 312) have been reported. Such stantial quantities of the aleurone layer, the layer yields significantly exceed yields of the highest which immediately surrounds the endosperm. yielding bread wheats (7245 kg/ha) grown under Another possibility suggested by Nelson (I 969) closely similar conditions. is to increase the production of a particular amino Trials are now in progress at CIMMYT to study acid above the level required for protein synthesis the response of promising triticale genotypes to by a mechanism studied by Vogel and Vogel (1967) nitrogen fertilizer under a variety of soil and en­ in microorganisms. Enzyme systems present in vironmental conditions. The first results, which will mutants of certain microorganisms appear to include grain and straw yields, kernel characteris­ behave abnormally giving rise to an unusually tics and protein content, will be available in the high production of a particular amino acid in free near future. form. No mutation demonstrating a similar effect From India come reports of triticale yielding has as yet been found in cereals (Nelson 1969). well at high altitudes and on acid soils, and in Ethiopia five triticale varieties are reported to have performed better than eight durum wheats at five Environmental and Agronomic Factors locations. Triticale nurseries are established in Every plant is the outcome of the interactions 52 countries covering a wide range of soil and which take place between its genotype and the environmental conditions under both rain fed and environment in which it is grown. Manipulation irrigated conditions, the rain fed areas embracing of this environment by means of the agronomic almost every known level of precipitation. practices adopted can influence the productivity, In addition, triticale appears consistently to the health and the biochemical composition of the display protein contents comparable to the best plant. bread wheats together with a biologically superior It is now well established that the quantity of amino acid composition. protein in a cereal is genetically controlled Consequently, though much more research is (Haun old et al. I 962a,b; Stuber et al. 1962; needed, one can feel extremely optimistic about Johnson et al. 1963, 1967, 1968a,b, 1970; Lonn­ the future of triticale as a valuable new source of quist 1969). However the fertility of the soil in calories and protein for many people. HULSE/LAING: PROTEINS OF TRITICALE 45

Triticale and grown during the winter season of 1969-70 (Sethi and Singh 1972). The strains differed Genetic and Varietal Effects significantly for all the characters studied. The estimates of correlation coefficients indicated that TOT AL PROTEIN the number of spikes per plant was the only Chen and Bushuk (1970c), from a comparison effective yield-contributing character. Regression of the disc electrophoretic patterns for the analysis indicated that spikes per plant, I 00-grain albumins, globulins, gliadins, and glutenins of weight, spike length, days to 70% heading, days one hexaploid triticale line (6A 190) and of its to maturity and plant height are important durum wheat (cv. Stewart 63) and rye (cv. Prolific) characters, contributing about 78% of the total parents showed that all the protein components variability for grain yield, if the selection is based of the triticale are present in the parents and con­ only on these characters. clude that the proteins of hexaploid triticale are The protein contents (N x 6.25) percent of simply inherited from its parents. Chen and four varieties of triticale grown at Fargo, North Bushuk (1970c) cite Yong and Unrau (1964) who, Dakota, USA in 1970 were: Rosner 18.2; 6TA203 using the same line of triticale but a slightly (FasGro 203) 17.9; 6TA204 (FasGro 204) 18.6; different extraction procedure, found new proteins 6T A209, 19.7. In 1971, the protein contents were: in the triticale. Rosner 15.3, 6TA203, 15.2; 6TA204, 15.3; Ellis (1971) using an electrophoretic system, 6TA209, 15.1 and of Trailblazer (a reselection of examined the gliadin fraction of a series of 6TA209) 15.8 (Busch and Wilkins, 1972). hexaploid triticales (PM 1514, I/ I, 2/ I, 3/ I, Sisodia ( 1973), reporting on research in India, AFPl/184) and cultivars of T. aestirum, T. durum states that considerable variability in protein and S. cereale obtained from the National content (N x 5. 7), 9 .8 to 16.3%, was observed Institute of Agricultural Botany and the Plant among different head progenies of a triticale Breeding Institute, Cambridge, England. He found variety. Normally in a routine breeding program, that the electrophoretic patterns were more no selection for protein content is practised and variable in triticale than in wheat and about as protein estimations are usually made at a later variable as in rye. Some triticale varieties resembled stage of development of a variety. Thus a variety wheat more closely than rye (e.g. AFPl/184) and otherwise homogeneous, is expected to be heter­ some resembled rye more closely than wheat ogeneous for characters which have not been (e.g. 3/1). selected. Accordingly, protein content in a variety Hristova and Baeva ( 1972) subjected two may represent an average estimate of high and low octaploid triticales to electrophoretic analysis. protein genotypes. Selection for high protein The triticales were AD-901, of which the parents content in an otherwise high yielding variety, were the wheat variety Norin-69 and rye Lozen-14, therefore, is expected to be effective. and AD-COC-3-2, of which the parents were the wheat Bezostaia I and the rye C2 . They found that certain of the protein components in the spectra AMINO ACID COMPOSITION of the parents were missing from the triticales Chen and Bushuk (I 970a) reported on the and further observed new components in the solubility characteristics and amino acid composi­ triticales. They did not find any simple form of tion of one line of triticale (6Al90) and of its inheritance of the protein components of the durum wheat (cv. Stewart 63) and rye (cv. Prolific) parents and therefore consider protein spectra of grown together in 1966 at the University of the triticales cannot be reported as a simple Manitoba with the hard red spring wheat, addition of the protein spectra of the parent Manitou. The samples were milled into flour on grains. the Buhler experimental mill after tempering Although protein quantity and quality are not overnight to 16.5% moisture. Standard analytical reported, it may be of interest to refer here to the methods of the American Association of Cereal studies carried out at the Plant Breeding Research Chemists ( 1962) were employed to characterize Substation, Bajaura, India on the genetic vari­ the flours (see Table 9). The amino acid composi­ ability, interrelationships and selection index of tion (by autoanalyzer) is given in Table 10. In 31 strains of triticale obtained from overseas general the amino acid composition of triticale 46 MONOGRAPH IDRC-02Ie

TABLE 9. Characteristics of triticale. rye. durum wheat and hard red spring wheat (data from Chen and Bushuk 1970a).

Durum Hard red Triticale Spring rye wheat spring wheat 6AI90 Prolific Stewart 63 Manitou

Grain IOOO-kernel weight. g 43.4 28.6 44.9 30.6 Moisture content. /~ 14.0 13.0 13.7 13.5 Flour 0 Yield. 0 I 14~·0 moisture has is) 58.9 59.6 67.7 71. 7 0 Ash content. 0 I 14° 0 moisture has is) 0.42 0.68 0.66 0.48

Moisture content. / 0 14.0 13.0 13.7 14.8

Protein content. '/'0 I14% moisture hasis) 9.8 9.9 12. I 13.6 was intermediate between its parent species. was reconstituted and 10 it was added. separately durum wheat and rye. and int urn. each of the seven pairs of chromosomes Riley and Ewan (1970) reported on studies of of the rye variety King II. Roman numerals were the amino acid composition of the T. aestirum assigned 10 the addition lines. Amino acid compo­ (bread wheat) variety Holdfast and the rye tion (by au10analyzer) is shown in Table 11. variety S. cereale King II. Chromosome addition The use of whole seeds instead of ground lines were constructed by simple backcross pro­ material. necessary because of the small quantity cedures using Holdfast as the recurrent parent. of material available. resulted in significant losses In the addition lines the genotype of Holdfast in both wheat and triticale of tyrosine. arginine. cystine and methionine. The effects on the amino TABLE 10. Amino acid composition of flours from acid content of grains of the addition 10 wheat of rye. triticale. durum wheat and hard red spring wheat the entire chromosome complement of rye and (111icromole.1· amino acid per milligram nitrogen in sample) some of the effects of each pair of rye chromosomes (Chen and Bushuk I 970a). in turn is shown in Table 11. Riley and Ewan point out that rye chromosome I increased HRS cystine and lysine by 10.7 and 8.7° 0 respectively. Rye Triticale Durum wheat Chromosome VII reduced the content of threonine

. by 10.4° 0 Aspartic acid 2.41 2.23 I. 89 1.47 Comparison of the values in Table 12 indicates Threonine I. 29 I. 34 I. 29 I. 20 that the contents in the triticale of threonine. Serine I. 84 2. 17 2.26 2.21 serine. glutamic acid. alanine. praline and tyro­ Glutamic acid 9. 77 II. 7 13.2 15.7 Pro line 6.26 6.44 6. 56 6.21 sine may be influenced by interaction among Glycine 2.40 2. 70 2.54 2.73 chromosomes. For the remaining amino acids Alanine 2. 15 2.08 I. 99 I. 84 there is no strong evidence that the contents of Valine 2. IO 2.20 2.21 2. II the triticale grain cannot be accounted for by Cystine 0. 52 0.70 0.60 0.59 simple additivity. Methionine" 0.68 0.79 0.78 0.65 Rye chromosome I is in homoeologous group 5 Isoleucine I. 51 I. 74 I. 87 I. 71 (Sears 1968). and Tsunewaki ( 1968) has reported Tyrosine 0.56 0.81 0.80 0.78 that the winter/spring habit difference in wheat is Phenylalanine 1.47 I. 58 I. 80 2. 10 primarily determined by the chromosomes of Ammonia 9.41 10.0 12. 5 13.6 group 5. Since Johnson et al. (I 968a) found that Lysine I. 12 1.00 0. 85 0. 82 eight of the ten varieties of wheat with the highest Histidine 0.65 0.79 0.83 0.85 lysine content also had winter habit. group 5 Arginine I. 23 I. 43 I. 13 1.20 Tryptophan not determined chromosomes may be of particular significance in N recovery. ",~ 76.2 84.5 87.8 91. 7 the control of lysine content in the grain. Con­ firmation of this would lead 10 a more rational "Including methionine oxide (approximate value). approach to breeding for higher lysine content. HULSE/LAING: PROTEINS OF TRITICALE 47

TABLE 11. Amino acid contents of the grains of wheat, rye, triticale and the seven addition lines with single pairs of rye chromosomes added to wheat (grams/JOO g of recorered anhydro acids) (Riley and Ewart 1970).

Addition lines

Triti- Amino acid Wheat II 111 IV v VI Vil ca le Rye

Aspartic acid 6.06 6. 13 5.74 5.54 6. 18 5.83 6.47 6. II 5.81 7.45 Threonine 3.66 3.59 3.34 3.56 3.58 3.83 3.40 3.28 3.62 4.06 Serine 5.66 5.51 5.40 5.54 5.54 5.55 5.54 5.53 5.46 5.50 Glutamic acid 37 .15 35.47 36.94 38.90 34.93 37.94 36.94 37.53 36.97 32.63 Pro line 12.94 13.62 14.12 13 .04 13. 31 13.00 14.47 13. 14 14.03 13.65 Glycine 4.89 4.86 4.60 4.58 5. II 5. 19 4.70 4.90 4.71 4.97 Alanine 4.27 4.19 3.97 4.04 4.28 4.05 4. 27 4.05 4. II 4.87 Valine 5.29 5.31 5. 14 5 .12 5.35 5.06 5.22 5. 15 5. II 5.73 Cystine I. 78 I. 97 1.80 1.84 I. 81 1.94 I. 89 1.80 1.80 I. 83 Methionine' I. 71 I. 82 1.66 I. 82 I. 71 I .67 I. 74 1.60 I .68 I. 85 lsoleucine 4.43 4.44 4. 50 4.32 4.43 4. 30 4. 30 4.39 4.32 4.44 Leucine 8.30 8.44 8. 17 8.00 8.33 8.01 8. 13 8.36 7.99 7.75 Tyrosine 2.55 2.67 2.98 2.70 3.00 2.58 2.56 2. 72 2.49 2.06 Phenylalanine 6.09 5.97 6.44 5.85 6.19 5.79 5.82 6. II 6. 12 6.27 Lysine 3.23 3.51 3. 17 3. 17 3.42 3.41 3. 17 3.21 3.22 4.24 Histidine 2.83 3.03 2.80 2.83 3.05 2.71 2.66 2.83 2.88 2.70 Arginine 5.37 5.69 5.34 5.32 6.00 5.76 4.94 5.43 5.80 6.30 Tryptophan not determined Total 116.21 116.22 116.11 116. 17 116. 22 116. 17 116. 22 116.14 116.12 116. 30 Protein (N x 5.7), % 15.4 16.7 19.9 16.4 19.7 15.5 18.2 19.0 17.3 18.2 Recovery.% 87.2 84.1 82.9 91.4 82.3 89.2 84.8 81. 5 83.6 86.3

'Including methionine sulphoxide.

Larter et al. (1968) reported on the protein Larter et al. (1968) do not discuss the improve­ content and amino acid composition of hexaploid ment in protein content and quality that may triticale tested in 1967 at the University of Mani­ result from selective breeding but point out that toba. The parentage of the lines tested was: the basic weakness in triticale rests in its repro­ ( T. durum (var. Ghiza) x S. cereale) x ( T. durum ductive system; an improvement in its genetic (var. Carleton) x S. cereale) x (T. persicum x S. stability will lead to increased fertility and yield. cereale) x (a Triticum x Secale introduction). The Their experience in improving kernel type has overall mean yield of the best triticale was about shown that in transmitting genes with favourable equal to the best bread wheat ( T. aestil'Um var. characteristics, some undesirable characteristics Manitou). The percentage protein and amino may accompany the favourable ones. For example, acid composition (except tryptophan) are given T. timopheeri and Secale montanum transmit genes in Table 13. The analytical methods are not stated. for desirable kernel characteristics but T. The amino acid balance of triticale compares timopheeri also transmits male sterility and S. favourably with that of wheat. The gradual decline montanum transmits extreme lateness and chro­ in protein level which the Manitoba triticale lines mosome anomalies. Since the Larter et al. paper have shown over the years is probably the result was published considerable improvement in ofa gradual improvement in kernel characteristics. fertility and kernel characteristics has taken The original, wide crosses of durum wheat and rye place (CIMMYT 1970-71, Zillinsky 1973). and of bread wheat and rye, produced a high Tkachuk and Irvine (1969) determined the proportion of badly shrivelled grains which, not amino acid composition by autoanalyzer of surprisingly, tended to be high in protein expressed several cereals, including triticale grown in 1966 as percent dry matter (Zillinsky and Borlaug by the Rosner Research Group, University of 1971a). Manitoba, the protein content of which, on an 48 MONOGRAPH IDRC-02le

TABLE 12. Effects on the amino acid content of the TABLE 13. Percentage protein. 1000-grain weight and grains of the addition to wheat of the entire chromo­ amino acid composition oftriticale in comparison with some complement of rye and the sum of the effects of bread wheat. 1967 (Larter et al. 1968). each pair of rye chromosomes in turn (Riley and Ewart 1970). T. aestivum Triticale (rnr. Manitou) (mean 6 strains) Sum of effects of 0 individual rye Protein (drr hasis), 0 12 2 12 9 Entire rye genome chromosomes 1000-g rain weight, g 28. 1 37.3 0 Amino acid ( Triticale-1rheat) (Addn. line-u·heat) Amino acids, 0 Aspartic acid 4.3 5.7 Aspartic acid -0.25 -0.42 Threonine 2.4 3.2 Threonine -0.04 -1.04 Serine 3.5 4.0 Serine -0.20 -1.01 Glutamic acid 32.5 27.2 Glutamic acid -0.18 -1.85 Pro line 10.6 9.5 Pro line 1.09 4.12 Glycine 3.9 4.4 Glycine -0.18 -0.29 Alanine 3. 1 4.2 Alanine -0.16 -1.04 Valine 4.1 4.8 Valine -0.18 -0.68 Cystine I. 9 2.0 Cystine 0.02 0.59 Methionine 1. 8 1 .2 Methionine -0.03 0.05 Isoleucine 3.4 3.7 Isoleucine -0. 11 -0.33 Leucine 6.5 6.8 Leucine -0.31 -0.66 Tyrosine 2 .1 2.4 Tyrosine -0.06 I. 36 Phenylalanine 4.6 4.8

Phenylalanine 0.03 -0.46 NH3 6.0 6.0 Lysine -0.01 0.45 Lysine 2.6 2.9 Histidine 0.05 0.10 Histidine 2.4 2.4 Arginine 0.43 0. 89 Arginine 4.5 5.2 Tryptophan not determined

as-is basis, was 15.6% (N x 5. 7), moisture 13.1 / 0 • 1966--69. These are given in Table 16, in com­ The amino acid composition is presented in parison with the Hungarian Official Standards for Table 14 in grams amino acid per I 00 g sample barley, wheat and rye. Table 17 gives crude protein nitrogen. The results for Manitou wheat (protein (N x 6.25) and the lysine, methionine and cystine 15.4%, moisture 8.1 %), Selkirk wheat (protein content (by autoanalyzer) as percent of grain of 15.0%, moisture 8.1 %) composites, grown in selected strains of these triticales. Protein con­ Western Canada in 1966, and for a dark rye flour tents ranged from 11.3 to 18.5% with an average obtained commercially (protein 13.5%, moisture of 13.2% at moisture levels ranging from I 0.0 8.8%) are also given. to 13.3% with an average of 12%. Among the Kurelec et al. (1968) presented data (see Table selected strains, protein varied from 11 .06 to 15) on the protein (N x 6.25) and amino acid 18.0% (13% moisture basis) and lysine, as percent composition of three strains of triticale designated of the grain, ranged from 0.20 to 0.67. 30-66, 57-66 and 64-66 compared with Bankuti Villegas et al. (1970) determined, by auto­ (Hungarian), San Pastore (Italian), Bezostaia 4 analyzer, the lysine content of the protein of a (Russian), Etoile (French) feed wheats and rye considerable number of samples of hard red grown under the same conditions in Hungary. spring wheat, durum wheat, and other wheat Among the wheats the highest content of lysine as species; of rye, grown under differing conditions; percent in the protein, was found in Bezostaia and of 25 varieties of triticale developed at the (3.6%). The rye contained 4.3% lysine and the University of Manitoba, and grown at Sonora, triticales averaged 2. 7%. Mexico in 1965-66. Gundel et al. (1970) reporting from the same Nitrogen was determined by micro-Kjeldahl, institute as Kurelec et al. (1968) gave details of the the conversion factors used being 5.7 for wheat average chemical composition of strains of hexa­ and triticale and 6.25 for rye. Values for protein ploid triticales raised in Hungary in the years are reported on a 14% moisture basis and those HULSE/LAING: PROTEINS OF TRITICALE 49

TABLE 14. Amino acid composition of triticale, Manitou and Selkirk wheat, and dark rye flour (in grams amino acid per JOO g sample N) (data from Tkachuk and Irvine 1969).

Selkirk wheat Manitou wheat Dark Triticale composite composite rye 1966 1966 1966 flour

Tryptophan 9.90 9.55 9.20 7. 57 Lysine 19.00 14.50 14.80 18.10 Histidine 15.50 13.80 14.30 13 .10 Ammonia 18.60 22.30 22.10 18.20 Arginine 30.50 24.90 24.40 26.20 Aspartic acid 36.90 29.20 30.90 40.40 Threonine 19.60 17.30 17.20 20.90 Serine 28.50 31.20 29.70 26.80 Glutamic acid 193.00 207.00 207.00 172.00 Proline 66.90 69.60 67.50 65.00 Glycine 24.70 23.50 23.90 22.70 Alanine 22.70 20.40 21.20 23.20 Cystine + Cysteine 17.40 16.20 15 .10 14.30 Valine 31.30 27.90 27.80 30.60 Methionine 12.00 10.50 10.60 07.31 Isoleucine 25.90 23.90 21.80 22.80 Leucine 42.00 42.00 41.30 37.40 Tyrosine 14.50 16.70 17.40 11.80 Phenylalanine 29.70 29.70 30.70 28.00 N recovery, % 95.30 96.40 95.80 89.60

TABLE 15. Protein (N x 6.25. 14% moisture basis) and amino acid content (as percent protein) of Hungarian grown triticales, wheats and rye (data from Kurelec et al. 1968).

Wheat Triticale Bankuti San Pastore Bezostaia 4 Etoile 30-66 57-66 64-66 Hungarian Italian Russian French Rye

Protein (N x 6.25), % 13.8 12.7 14.2 16.6 13.7 14.0 12.4 9.4 Amino acids, % protein Leucine 8.0 8.0 7.2 7.7 8.6 7.3 Isoleucine 3.5 3.4 2.9 5.4 4.8 3.8 Phenylanine 4.8 3.8 4.4 6.0 3.3 3.3 5.0 4.5 Methionine I. 5 I. 3 I. I 1.4 1.4 I.I Valine 5.5 5.4 4.8 4.9 6.5 4.6 Threonine 4.9 4.4 3.9 3.7 2.7 3.0 4.5 3.8 Glycine 7.6 7.0 8. I 7.7 5.3 4.9 4.3 6.6 Glutamic acid 27.2 28.2 34.5 23.9 27.3 26.6 20.5 15. I Arginine 3.3 3.0 5. I 3.6 4.0 4.4 4.3 5.7 Lysine 2.7 2.6 2.8 2.5 2.5 3.6 2.7 4.3 Histidine I. 8 I. 9 2.1 2.4 2.5 2.8 2.2 2.4 Cysteine 1.4 I. 5 1.6 1.0 1.0 50 MONOGRAPH IDRC-02le

TABLE 16. Average percent composition (ranges in parentheses) of Hungarian triticales analyzed in 1966-69, compared with Hungarian standards for barley, wheat and rye (data from Gundel et al. 1970).

MZ6830-66 (Hungarian Standard)

Triticale Barley Wheat Rye

Dry matter 87.0 87.0 87.0 87.0 (86. 7-90.0) Moisture 12.0 13.0 13.0 13.0 (10.0-13.3) Crude protein (N x 6.25) 13.2 10.2 12.2 11.6 (11.3-18.5) Crude fat 1.8 1.8 1.9 I. 7 (1.3-2.7) Crude fibre 2.4 3.7 1.9 1.9 (1.4-3.8) N-free extracts 67.8 68.8 69.3 69.8 (65. 0-80. 0) Crude ash 1.8 2.5 I. 7 2.0 (1.4-2.4) Starch 50.7 (46. 5-54 .4)

TABLE 17. Protein, lysine, methionine and cystine content of various strains of Hungarian triticale (I 3% moisture basis) (data from Gundel et al. 1970).

Protein Sample (N x 6.25) Lysine Methionine Cystine Number (%) percent of grain

20/68 17.57 0.36 0.11 0.19 30/68 18.00 0.67 0.14 0.43 57/68 11.06 0.27 0.10 0.16 57/68 11. 79 0.31 0.11 0.20 57/69 14.81 0.37 0.11 0.12 64/St 11.59 0.41 0.09 0.16 64/68 12.91 0.30 0.14 0.15 64/68 11.17 0.20 0.10 0.17 64/69 16.47 0. 33 0.11 0. 32

for lysine are reported on a dry weight basis. The Fargo) to 18.8% (Justin at Minot). In Mexico, results for triticale are given in Table 18. The the protein content ranged from 10.6% (Crim) to lysine content in the protein was significantly 16. 7% (Justin). Lysine in the protein ranged from inversely correlated with the protein content of 2.15 g/16 g N (Justin in Mexico) to 2.77 g/16 g N triticale (r = -0.50). (Lerma Rojo in Mexico). There was a low nega­ The hard red spring wheat varieties, Selkirk, tive correlation (r = -0.4) for lysine in the Justin, Chris, Manitou and Crim were grown at protein vs. protein content for the higher protein two locations in North Dakota, and Selkirk, content wheats grown in North Dakota and a Justin, Chris and Crim with the soft wheat varieties significant high negative correlation (r = -0.68) Lerma Rojo, Narino and 8156, were grown in for the lower protein content wheats grown in Mexico. The protein content of the wheats grown Mexico. The overall correlation was -0.49 and in North Dakota ranged from 15.0% (Selkirk at was significant. HULSE/LAING: PROTEINS OF TRITICALE SI

TABLE 18. Lysine content of triticales (data from Villegas et al. 1970) (Villegas 1972-73 data added in proof).

Average Average lysine Villegas 1972-73 protein (N x 5.7 In protein Lysine 14% moisture (g/16 g N In sample Lab Protein % Variety or Cross' basis)(%) dry weight basis) (%) No. % protein

1593A x 1620 14.7 3.08 0.577 5731 13.0 4. 35 15938 15. I 3.08 0.594 5734 13.2 4.27 1593C 16.4 3.06 0.641 5735 13.9 3. 72 15930 14.3 3.26 0.589 5742 13.0 4.17 1594A 16.2 2.98 0.615 5744 13.0 3. 72 1594A x 1601 15.8 3. I I 0.627 5746 12.5 4.28 1594A x 1613 16.6 3.35 0. 711 5792 12.4 3.81 1594A x 1628A 14.5 3.02 0.559 5793 12.3 4.04 1605 14.6 2.89 0.539 5796 12.2 4.05 16098 x 1636 16.6 2.92 0.618 5800 12.4 3.78 1636A 13.9 3.25 0. 578 5801 12.3 3.83 1636A x 1614 15.0 3.08 0.590 5804 12.6 4.05 1636C 14.3 3.02 0.556 5805 13.3 3.87 1636C x 1642 14.5 3.04 0.563 5807 12.4 4.05 1637C 16.0 2.92 0.597 5819 13.4 3.78 1641A 12.8 3. 12 0.507 5821 12.2 4. 13 16418 13. I 3.04 0.509 11452 16.3 2.98 1641D 13.8 3.21 0.565 11453 11. 2 3.56 6A250 x 6Al90b 16. l 2.73 0. 557 6A250 x 6Al91' 13.3 2.94 0.496 My 64 x Triticale 17.0 2. 75 0.595 Triticale S-112 13.8 2. 72 0.479 Triticale 6913 18.5 2.71 0.639 Triticale IOO-C-132-1 14.7 2.74 0.563 6A250(6A66.12 x 6A20) 15.0 2.82 0.540

'Average lysine and protein content of selections from each variety or cross. bOne selection contained the low of 2.32 g lysine per 16 g Nin the protein. 'One selection contained the high of 3.42 g lysine per 16 g Nin the protein.

Five varieties of durum wheat, Mindum, Wells, protein was lower than the mean of 2. 71 g lysine/ Lakota, Stewart 63 and Leeds were grown at three 16 g N in the protein reported by Lawrence et al. locations in North Dakota, and 23 lines were ( 1958) for six durum wheat varieties with an grown in Mexico. At North Dakota, protein average protein content of 13%.

varied from 13.8% (Mindum at Fargo) to 19.9/0 Altogether 125 varieties or species of rye were (Leeds at Edgeley). Lysine in grams/ 16 g N in the analyzed by Villegas et al. (1970); the value for protein varied from 1.84 g (Leeds at Edgeley) to 20 of these are presented in Table 19. Lysine 2.45 g (Mindum at Edgeley) and the negative content in the protein correlated significantly correlation between lysine in the protein and pro­ with the protein content (r = -0.69). It will be tein content was highly significant (r = -0.77). observed from Table 19 however that the varieties In Mexico, protein varied from 9.5% (RD! 76-7 A) Explorer, USDA Pl 168178, Rye Gator, Detenicke to 17.1%(RD!0!-2A). Lysine in the protein varied and Secale montanum-23-282 were relatively high from 2.17g (RDI82-llB) to 3.!0g (RDI76-7A) in both protein and lysine. but the correlation with protein was not significant Since the lysine varies with the protein content, (r = - 0.33). However, the overall correlation the regression equation for lysine in protein vs. coefficient for durum wheat was -0.65 and protein content was used first to estimate the significant. The overall mean protein content was overall lysine content at 13.S and 17.2% protein 15.3%. The mean of 2.39g lysine/16gN in the and secondly, the variability of lysine content for 52 MONOGRAPH IDRC-02le

TABLE 19. Lysine content of rye species and varieties (Villegas et al. 1970).

Lysine

Protein In protein In sample Variety and/or Species (".,) (g/16 g N) C' o>

Antelope 11.4 3.33 0.443 Carsten 9.2 3.65 0.392 Secale dalmaticum CPI 22755 16.5 3.23 0.621 Explorer' 14. I 3.30 0. 542 USDA Pl 168178' 14.6 3.22 0.549 USDA Pl 227870 10.5 3.71 0.453 Rye Gator' 17.3 3.21 0.649 Prolific Spring 14.9 3.25 0.563 Ba Ibo 11 .2 3.63 0.474 Detenicke' 16.0 3.37 0.628 Secale cereale 5-SC-18 12.8 3.22 0.482 Rye Korean I 15.3 3.05 0.544 Dominant 11.0 3.80 0.489 Volyanko 11.9 2.91 0.400 Afganistan Winter Rye 10.4 3.35 0.405 Canadian Spring Rye 9.2 3.71 0. 397 Secale montanum' 23-282 16.9 3. IO 0.611 Secale segetale 23709 17.2 3.02 0.604 Abruzzi A 9.9 2.55 0.292 Maia barroso (5053) 10.6 4.26 0.524

'Ryes with relatively high lysine and protein. each cereal examined. The results are given m protein content of the triticale is higher than the Tables 20 and 21 . wheat and rye, but the lysine and threonine Munck (1972) reported on the amino acid contents, expressed as grams amino acid/16 g N analyses (by ion exchange) of different varieties are lower in triticale than in wheat and rye. of wheat, rye and octapoloid triticales grown at Ruckman et al. (1973) compared the hard red two locations in Sweden. From the results pre­ spring wheat INIA 66, the white spring wheat sented in Table 22, it will be seen that the mean Siete Cerros 66, and the amber durum spring wheat Oviachic 65, all short varieties, with the TABLE 20. Estimated lysine content of wheat. rye. and triticales 6TA204 (Jenkins Foundation for triticale at 13.5 and 17 2°,, protein (dry 1reigh1 hasis) Research, Salinas, California); Rosner (University (Villegas et al. 1970). Manitoba) and T-1324 (CIMMYT), all tall varieties, in field experiments at El Centro, Five Lysine per 16 g N in protein Points, Davis and Tulelake, California, using

13.5",, 17.2"~ TABLE 21. Variability of the estimated lysine content

protein protein (al 15° 0 pro1ein /ere[) in wheat. rye and triticale Cereal (g) (g) (Villegas et al. 1970).

Spring wheat 2.49 2.36 Standard deviation Durum wheat 2.51 2.26 Cereal (as lysine per 16 g Nin pro!ein) Rye 3. 30(3. 39)' 3 .01(3. I I)' Triticale 3.06 2.81 Spring wheat 0. 13 Durum wheat 0.17 'Estimated value when a N-to-protein factor of 5. 73 Rye 0.24 is used [5. 73 is average reported by Jones ( 1931) and Triticale 0.21 Tkachuk (1969b)]. HULSE/LAING: PROTEINS OF TRITICALE 53

TABLE 11. Protein (N x 5.7) and content of nine amino acids in rye, octaploid triticale and wheat grown at two locations in Sweden in 1964 as grams amino acid/ 16 g, N (ranges in parentheses) (data from Munck 1972).

Winter Winter Winter Winter Spring rye triticale wheat wheat wheat (Svalof) (Lund) (Lund) (Svalof) (Svalof)

0 Protein. 0 10.8 14.8 II. 7 13. I 12.9 (9.6-12.4) ( 13. 5-16. I) (11.1-17.2) (IO. 515. 3) Amino acids Lysine 3.72 2.78 2.95 2.85 2.87 (3.3 4.0) (2.6-3.2) (2.5-3.0) (2.6-3.2) Arginine 5. 03 4. 30 4.74 4.67 4.66 (4. 6-5. 4) (4.0-4. 7) (4.4-5. I) (4.4-5.1) Histidine 2.30 2. II 2.30 2.28 2.24 (2. I 2.4) (I. 9-2.4) (2. 2-2. 4) (2. 1-2.4) Methionine I. 57 1.44 I. 52 I. 57 1.49 (without oxidation) (I .4-1. 7) (I .2-1.6) (I. 5-1. 7) (I. 3-1. 7) Threonine 3.58 3 .09 3. II 3. IO 3. 15 (3.4-3. 7) (2.9-3.3) (3.0-3.2) (2.8-3.3) Valine 4.88 4.42 4.46 4.53 4.42 (4.6 5. I) (4.2-4. 7) (4.4-4. 7) (4.0-4. 7) lsoleucine 3.56 3.46 3.48 3.61 3.55 (3. 5- 3. 8) (3. 2-3. 6) (3.4-3.9) (3.6-3. 7) Leucine 6.47 6.57 6.83 6.90 6.85 (6.1-6. 9) (6.3-6.9) (6.7 7.3) (6.4-7. I) Phenylalanine 4.82 4.64 4.41 4.61 4.60 (4.6 4.9) (4.4-4.9) (4.4-5.0) (4.4-4.8) ------·· chemical fertilization described as that employed triticales, T-1324 was unstable for grain, protein for optimum production at each location. On and lysine yields and Rosner for lysine yield; average, the triticales had higher whole-grain 6TA204 showed good stability. Among the wheats, protein content (Kjeldahl N x 5.7) than the ISIA 66 was unstable for grain. protein and lysine wheats (16.3 vs. 14.3% on a dry weight basis), yields, Siete Cerros 66 was unstable for grain and higher lysine (by gas chromatography) in the protein yields; Oviachic 65 was extremely stable. grain (0.428 vs. 0.307'1,,) and in the protein (2.63 vs. 2.14%). Mean yields of the triticales were lower at BIOLOGICAL QUALITY all locations; about 71% of the wheats. The best Studies on Human Adults Rosner triticale yield of triticale was from 6TA204 at Tulelake grown in Manitoba, Canada and wheat (a com­ which has longer days and cooler conditions than posite of high-protein Atlas 66 x Comanche lines the other locations; the worst was from Rosner, grown in Nebraska, USA) were compared, at which does not perform well in California, at two levels of protein intake, in nitrogen balance El Centro. studies with young human adults by Kies and Fox Grain protein contents did not vary significantly (1970c). among locations. Protein yields of the triticales, The subjects were five males and four females on a dry weight basis, were significantly lower than and the period of the study was 33 days, consisting the wheats at all locations (540 lb vs. 6 75 lb/acre; of an introductory 3-day nitrogen-depletion 605 vs. 756 kg/ha), but lysine yields were com­ period, two 5-day adjustment periods and four parable ( 14.2 vs. 14.5 lb/acre; 15.9 vs. 16.2 kg/ha). experimental periods of 5 days each. The experi­ Production of grain showed significant variety mental periods were arranged at random for each x location interaction and this was reflected in the subject. During the adjustment and experimental protein and lysine yields per hectare. Wrickc's periods nitrogen (N) intake was maintained at ( 1962) method was used to determine varietal 4.8 or 6.8 g N per day respectively, 4.0 and 6.0 g N stability in relation to environment. Among the being provided by wheat or triticale, 0.8 g N from 54 MONOGRAPH IDRC-02le the basal diet. The wheat and triticale grains were and triticale flours were fed as baked yeast raised whole ground in a hammer mill and were fed in rolls. The amino acid supplements were L-lysine the form of baked, yeast raised rolls taken in equal (0.960 g per day wheat study; 0.891 g per day amounts at each of the three daily meals. The triticale study), L-methionine (1.152 g per day caloric intake for each subject was kept constant wheat study, 1.092 g per day triticale study) during the experimental periods at the level L-tryptophan (0.212 g per day wheat study, 0.236 required for weight maintenance. g per day triticale study). During one experimental In nitrogen balance studies, a decrease in body period no amino acid supplements were given nitrogen loss is generally interpreted as indicative (negative control) and in another a combination of improved protein utilization. The results of all three was given (positive control). showed that at the lower level of protein intake, 4.0 Protein was evaluated primarily by a nitrogen­ g N/day, the average nitrogen balance was -0.62 balance technique and by other laboratory tests, g N/day for wheat and -0.44 g N/day for triticale, as described by Kies and Fox (l 970c). The lysine a difference of 0.18 g N/day in favour of triticale, and methionine composition of the wheat and significant at the 5% level. At the higher level of triticale were determined by autoanalyzer; trypto­ protein intake, 6.0 g N/day, subjects on both the phan was estimated from reference tables. wheat and triticale diets showed a significantly Under the experimental conditions of both better nitrogen retention than when receiving the studies, lysine was shown to be the first limiting 4 g N level diets. The average nitrogen balance amino acid. The average increment in nitrogen was -0.16 g N/day, for wheat and+ 0.01 g N/day retention between lysine-supplemented and un­ for triticale, a difference of0.17 g N/day in favour supplemented wheat diets was 0.73 g N per day, of triticale, significant at the 1% level. and in the triticale diets 0.54 g N per day, both

Two subjects did not conform to the response significant at the 1°10 level of probability. pattern. Differences in individual response are Rat, Mouse, and Vole Assays Knipfel (1969) not, however, unusual in nitrogen balance studies. compared a sample of Rosner triticale with wheat Female subjects tended to retain more nitrogen and rye samples purchased locally, in diets fed to than males possibly because the latter engaged in male weanling rats. The diets contained 10% a higher level of strenuous exercise during the protein from casein, triticale, wheat, rye or equal­ experimental period which may have affected part mixtures of casein plus wheat, casein plus protein as well as caloric need. triticale or casein plus rye. The protein contents Kies and Fox (l 970b) also conducted two studies were: casein 86.3% (N x 6.25); wheat 13.5% with human volunteers to determine the first (N x 5.7); triticale 16.1% (N x 5.7); rye 12.6% limiting amino acid in ground whole triticale grain (N x 5.7). Food and drink were provided ad (Rosner grown in Manitoba, Canada) and in libitum. The amino acid composition (by auto­ whole ground wheat (a composite of high-protein analyzer) of the diets, except forcystine and trypto­ Atlas 66 x Comanche grown in Nebraska, USA). phan, is shown in Table 23. The Protein Efficiency Nineteen young adults took part in two studies Ratios (PER), adjusted to a value of 2.50 for each 33 days in length. In the wheat study seven casein, are given in Table 24. The PER of Rosner males and three females took part and in the triticale was equal to the PER of rye, with the PER triticale study six males and three females took of wheat significantly lower. part. Each study had a 3-day nitrogen-depletion Blood samples from each dietary group were period, a 5-day nitrogen adjustment period and pooled and analyzed by microbiological assay for five experimental periods of five days each. The free lysine, methionine and threonine. The results experimental periods were arranged at random indicate that lysine was markedly limiting in the for each subject. After the nitrogen-depletion triticale and the wheat, but less so in the rye. period, nitrogen intake was maintained at 5.0 g N Subsequent limiting amino acids were not deter­ per subject per day, 4.0 g N from wheat or triticale mined. Knipfel (1969) considers that the superi­ plus 0.68 g N from the basal diet and systematically ority of triticale over wheat may be attributed to variable amounts of nitrogen from the amino acid its higher content of lysine and sulphur amino supplements. Urea was used to maintain the diets acids. isonitrogenous at 5.0 g N intake level. Caloric in­ In isonitrogenous diets fed to weanling rats, take was kept constant for each subject at the Shimada and Cline ( 1972) compared two unidenti­ level required for weight maintenance. The wheat fied varieties oftriticale, one grown in Indiana and HULSE/LAING: PROTEINS OF TRITICALE 55

TABLE 23. Amino acid composition of test diets calculated from amino acid analyses of individual protein sources (Knipfel 1969).

Diets

(CJ 5~/~ (E) (G) Tri ti- 5% 5% (A) (B) cale; (D) Casein; (F) Casein; 100,0 10% 5% 10% 5% 10% 5% Amino Acid Casein Triticale Casein Wheat Wheat Rye Rye

percent o(diet Aspartic acid 0.68 0.51 0.59 0.49 0. 59 0.74 0.71 Threonine 0.37 0.29 0. 33 0.32 0. 35 0. 33 0.35 Serine 0.46 0.40 0.43 0.41 0.44 0.37 0.42 Glutamic acid 2.30 3. 19 2.75 3.71 3.00 3. 12 2.71 Pro line I .II I. 21 1.16 0.91 1.01 0.90 1.01 Glycine 0.19 0.42 0.30 0. 39 0.29 0.43 0.31 Alanine 0.30 0.39 0. 35 0.36 0. 33 0.44 0.37 Valine 0.63 0.47 0. 55 0.48 0. 55 0.45 0. 54 Methionine 0.26 0. 15 0.21 0.10 0.18 0.13 0.20 Isoleucine 0.54 0.41 0.48 0.40 0.47 0.37 0.46 Leucine 0.92 0.69 0.80 0.65 0.79 0.56 0.74 Tyrosine 0.50 0.32 0.41 0. 36 0.43 0.27 0.39 Phenylalanine 0.49 0.49 0.49 0.48 0.49 0.42 0.45 Lysine 0.71 0. 33 0. 52 0.29 0.50 0.47 0.59 Histidine 0.27 0.24 0.26 0.25 0.26 0.26 0.27 Arginine 0.33 0.49 0.41 0.51 0.42 0.52 0.42

TABLE 24. Performance data of rats fed casein, triticale, wheat and rye diets (Knipfel 1969).

Diets*

(CJ 5% (E) (G) Tri ti- 5% 5% cale; Casein; Casein; (A) (B) + (D) + (F) +

10% I 0'/0 5% 10% 5% 10% 5% Criteria Casein Triticale Casein Wheat Wheat Rye Rye

Weight gain, g 63.7a 31. 8bc 67.2a 21. Sc 62.6a 37. 8b 73. la Feed intake, g 220.6a 177 .3b 234.0a 180.3b 240.3a 203. Ob 239.6a PER 2.50a I. 55c 2.49ab l .03d 2.25b I .6lc 2.64a

*Means with same letter(s) not significantly (P < 0.05) different. the other in Texas, with maize and maize plus basis. When the triticales provided the sole soya. The triticale varieties were also compared source of protein, at levels of 10% or 15%, and with maize alone on an equal weight in diet basis. were supplemented with amino acids, lysine was As judged by weight gain and feed: gain ratio, both found to be the first, and threonine the second, triticales were equal to maize on an isonitrogenous limiting amino acid. Methionine appeared to be basis and superior to maize on an equal weight third limiting. The Texas grown triticale was 56 MONOGRAPH IDRC-021e

TABLE 25. Biological evaluation of triticale samples from CIMMYT with mice (mean ra/ues offire mice) (data from P. J. Mattern 1972 personal communication).

Protein N x 5.7 (dry weight Weight Feed Feed Triticale basis) gain consumption efficiency Sample No. % (g) (g) ratio

A71-5403 16.1 11.54 112. 88 9.78 A71-5404 17.0 9.96 107.30 10.77

TABLE 26. Performance of growing rats on diets of Rosner triticale, Neepawa wheat and Cougar rye, fed to provide 10% and 7% protein in the diet, over 28 days (unpublished data from McDonald and Larter 1972).

Average daily Average daily Protein index gain feed intake (g u·t. gain/ 0---28 days 0---28 days g protein (g) (g) intake)

10% protein level Casein 3.64 13.95 2.51 Triticale I. 71 13.23 1.29 Wheat I. 11 9.90 1. 13 Rye I. 52 12.51 1.29 7% protein level Casein 2.60 16.18 2.20 Triticale 1.61 15.02 1.47 Wheat 1.14 12.31 1.26 Rye 0.93 13.70 1.01 judged to be of higher protein quality than the one relatively poor results which could not be com­ grown in Indiana. Weber and Reid (1972) have, pletely explained by differences in protein quantity however, reported briefly that in feeding trials and amino acid composition. with young mice, the first limiting amino acid The College of Agriculture at the University of for nineteen Mexican dwarf wheat varieties and Nebraska, Lincoln, Nebraska have also conducted for three (unidentified) triticale varieties was mice feeding trials on samples of triticale from methionine. The second limiting amino acid was CIMMYT. P. J. Mattern (1972 personal com­ lysine in the wheat and threonine in the triticale. munication) reported the results shown in Table The gains in body weight were about the same for 25 and also found substantial differences in feed­ all the wheat and triticale samples studied in ing trials with mice on wheat samples of the same Weber and Reid's"experiments but the liver GOT protein content and consumption. (glutamic oxaloacetic transaminase) activity and McDonald and Larter (1972 unpublished data) carcass retention of nitrogen differed though in compared Rosner triticale, Neepawa wheat and what manner is not stated. Cougar rye in feeding trials using rat, mouse and Octaploid triticale, protein (N x 6.25) 14.8%, meadow vole (Microtus pennsylvanicus). The winter rye, protein I 0.8%, and winter wheat, cereal (or casein in the control diet) was the sole protein 13%, were included in feeding trials with source of nitrogen and was fed to give a level of mice in a 20-day experiment (Munck 1972). The 10% protein (N x 6.25) in one series and 7% dietary protein varied with the protein content of protein in a second series. In the first series, I 0% the cereals. The triticale diet gave good results, protein level, diets containing soybean meal, comparable to that of the wheat. The rye diet gave wheat gluten, horsebeans (Vicia faba) and oats HULSE/LAING: PROTEINS OF TRITICALE 57

TABLE 27. Performance of mice on diets of Rosner triticale, Neepawa wheat and Cougar rye fed to provide l 0°1,, and 7% protein in the diet, over 21 days (unpublished data from McDonald and Larter 1972).

Average daily Protein index Average daily feed intake (g 11·t. gain/ gain, 0-21 days 0-2 l days g protein (g) (g) intake)

10% protein level Casein 0.62 4.71 l. 27 Triticale 0.40 3.91 l .04 Wheat 0.40 3.82 l.06 Rye 0. 36 3.70 l.03 7% protein level Casein 0.33 4.58 l. 10 Triticale 0.41 4.76 l. 16 Wheat 0.40 4.95 l. 12 Rye 0. 34 4.40 l .16

TABLE 28. Performance of voles on diets of Rosner triticale, Neepawa wheat and Cougar rye fed to provide 10% and 7% protein in the diet, over 21 days (unpublished data from McDonald and Larter).

Average daily Protein index Average daily feed intake (g 11·t. gain/ gain, 0-2 l days 0-21 days g protein (g) (g) intake

10% protein level Casein 0.30 3.30 0.89 Triticale 0.38 3.20 l .26 Wheat 0.24 3.02 0.75 Rye 0.30 3. 18 l.04 7% protein level Casein 0.16 3.68 0.60 Triticale 0. 35 4.21 l .19 Wheat 0.30 4.12 0.98 Rye 0.29 4.10 l .08 were also compared. In the second series, 7% were of the order of 50% of the mean values, protein basis, only diets containing casein, wheat, whereas those for rats were of the order of 20% rye and triticale were included. of the means. The measurement of body weight When fed for 21 days on the twelve different and food consumption for casein, triticale, wheat protein sources, the voles showed very little and rye are presented in Table 26 for rats, in Table fluctuation in body weight gain (grams per day) 27 for mice and in Table 28 for voles. Further the results varying between 0.16 g and 0.38 g, the tests of ten days duration were carried out with lowest value being on casein and the highest on rats, mice and voles using two samples of wheat triticale. In comparable experiments with rats, and four samples of triticale provided by body weight gain ranged from 0.22 g per day on CIMMYT. The chemical analysis of the wheat wheat gluten to 3.64 g per day on casein. Triticale and triticale samples, including their resorcinol showed an average gain of 1.71 g per day and soy­ content, is given in Table 29 and the results of the bean meal 3.10 g per day. The important result was feeding trials on these samples in Table 30. The that the standard errors in the vole protein indices laboratory rats and mice rated the biological 58 MONOGRAPH IDRC-02Ie

TABLE 29. Chemical analyses on 1ri1icale and wheal ing to their biological quality. Several of the voles samples from ClMMYT (unpublished daia from consistently rated cereal grains higher than casein. McDonald and Lafler 1972). Consequently the results from the vole assays cannot be regarded as a reliable indication of Pro1ein comparative nutritional values. (air-dry Gundel et al. (1970) compared three hexaploid basis) Lysine Fibre Resorcinol Sample (g/JUUg) (g//6g N) (g/IUUg) (g/JUU g) triticales, designated 20/68, 57 /68 and 64/68 with Bezostaia wheat, B.40 barley and Kecskemeti rye Wheal all grown in Hungary, in nitrogen metabolism 4859 II. 8 2.40 2.8 0.067 trials with rats. The protein (N x 6.25) and 4860 12.4 2.38 2.4 0.083 moisture content of the grains with their bio­ logical value, apparent digestibility of the protein Trilicale 4861 15.9 2. 72 2.7 0.091 and NPU are shown in Table 31. The authors 4862 15.6 2.80 2.4 0.099 consider that triticale would be useful as animal 4863 17. l 2.78 2.5 0.089 feed in Hungary. Chick Feeding Trials A hexaploid triticale of protein 18.42% (N x 6.25) was investigated by Sell et al. (1962) as a component of rations for quality of the cereals in an order predictable from male chicks. The amino acid content of the triti­ their chemical composition and/or previous ex­ cale, determined chromatographically, is com­ perience, but the variability among the voles made pared with the average composition of a hard red it impossible to classify the cereal varieties accord- spring wheat in Table 32.

TABLE 30. Biological evalua1ion of 1ri1icale and wheal samples from ClMMYT wilh rals, mice and voles (unpublished dala from McDonald and Lafler 1972).

Diel

(A) (B) (C) (D) (E) (F) Wheal Wheal Trilicale Trilicale Trilicale Casein 4859 4860 4861 4862 4863

Rais (means of 8) Average daily gain, g 4. 36 2.23 2.41 3.06 3.24 3.01 ±0.80 ±0.52 ±0.45 ±0.48 ±0.64 ±0.40 Average daily feed, g 17.38 17.80 16.24 21 .07 17.30 16. 94 ± 1.67 ± 1.45 ± 1.37 ±4.82 ± 1.62 ± 1.41 Pro1ein index (g 1\'t 2.28 1. 32 1. 36 1 .62 1. 71 1.66 gain/g protein intake) ±0.31 ±0.26 ±0.26 ±0.17 ±0.22 ±0.20

Mice (means of 10) Average daily gain, g 0.89 0.70 0.73 0.71 0.78 0.85 ±0.14 ±0.00 ±0.00 ±0.10 ±0.10 ±0.10 Average daily feed, g 5.30 5.34 5.61 5.43 5.80 5.56 ± 1.07 ±0.52 ±0.46 ±0.64 ±0.92 ±0.52 Pro1ein index (g 11·t 1. 54 1. 26 1.19 1. 22 1. 25 1.40 gain/g protein intake) ±0.28 ±0.14 ±0 14 ±0.17 ±0.24 ±0.14

Voles (means of 8) Average daily gain, g 0.39 0. 57 0.59 0. 58 0.57 0.42 ±0.14 ±0.19 ±0.23 ±0.16 ±0.12 ±0.14 Average daily feed, g 2.67 2.99 3.28 3.08 3. 23 2.49 ±0.25 ±0.60 ±0. 71 ±0.33 ±0. 53 ±0.27 Pro1ein index (g wt 1 .37 1. 82 1.64 1. 76 1. 59 1. 53 gain/g protein intake) ±0.56 ±0.41 ±0.49 ±0.46 ±0.16 ±0.38 HULSE/LAING: PROTEINS OF TRITICALE 59

TABLE 31. Protein and moisture contents of Hungarian grown triticales. Bezostaia wheat, B.40 barley and Kecskemeti rye and their biological value. apparent digestibility and NPU for rats (data from Gundel et al. 1970).

Apparent Protein Biological digestibility N x 6.25 Moisture value of the protein NPU Grain (%) (o;,) (o~) (%) ('/o)

20/68 Triticale 15.30 11.04 71.49± I .09 74.15 61. 29 57 /68 Triticale 12.00 10.34 72.83 ±0. 91 74.55 65.95 64/68 Triticale 9.45 10.75 72.85± 1.80 72.85 64.82 Bezostaia wheat 10.60 8.70 70. 77±2.40 72.17 59.83 B.40 barley 11. IO 10.09 74.21 ±0.88 69.12 62.69 Kecskemeti rye 10.80 8.77 74.82±0.81 65.66 60.76

TABLE 32. Amino acid composition of the protein of 1958) values of the 45, 60 and 67% triticale and triticale compared to that of an average hard spring wheat rations were similar; that of the 30% wheat (data from Sell et al. 1962). triticale significantly higher. The 81 % triticale ration, which was isonitrogenous with the wheat Amino acid Triticale Wheat basal, gave a significantly lower F/G than the wheat basal. The method by which the ration was (percent a/protein based on made isonitrogenous is not stated but Sell et al. N x 6.25) Arginine 4.74 6. 15 suggest that the poorer results from the 81 % Glycine 3.78 6.92 triticale ration may have been caused by reducing Histidine 2.18 2.31 the soybean protein in the isonitrogenous ration lsoleucine 3.73 4.62 to a point where deficiency in lysine resulted. Leucine 6.23 7.69 In the second trial, the basal ration contained Lysine 6.07 3.84 58.5% barley (12.27% protein) and 30% soybean Methionine 1.60 I. 54 meal (44% protein). Hard spring wheat and Phenylalanine 4.47 5.38 triticale were substituted for the barley on a 3.08 Threonine 2.50 weight for weight basis. Weight gains and F/G Tryptophan I. 17 I. 54 were not significantly different but the data sug­ Valine 4.26 4.62 gested that triticale was equal to wheat and superior Protein.% 18.42 14.18 to barley on a weight for weight basis. In the third trial, the basal ration contained 90% triticale and 1.5% soybean meal (44% protein), a In the first trial, the basal ration contained 67.5% total of 17.5% protein. The basal ration was hard spring wheat (14.18% protein) and 23% supplemented with 0.15% glycine, 0.05% DL­ soybean meal (44% protein). Triticale was substi­ methionine and 0.10% L-lysine HCl in a complete tuted for corresponding quantities of wheat at factorial arrangement. L-lysine HCI significantly 30, 45, 60 and 67% of the ration. A further ration improved weight gain and F/G. There was no which was isonitrogenous to the wheat basal response to glycine or DL-methionine. ration but contained 81 % triticale was included. In the fourth trial, the basal ration contained The rations containing from 30 to 67% triticale 76% triticale and 13.5% soybean meal (44% supported weight gains equivalent to the wheat protein), a total of 21 % protein. The test rations basal ration. The 81 % isonitrogenous triticale were: (A) basal + 0.1 % methionine, (B) basal + ration gave significantly lower weight gains. The 0.1 % lysine, (C) basal + 0.1 % methionine + 0.1 % efficiency of feed utilization and feed: gain ratio lysine, (D) basal + 0.2% additional protein, and (F/G) by chicks receiving 30 and 45% triticale (E) an isonitrogenous ration based on hard spring was significantly better than those receiving the wheat. Chick growth improved significantly over wheat, 60 and 67% triticale rations. The ration the basal ration only when both 0.1 % methionine metabolizable energy (ME) (Hill and Anderson and 0.1 % lysine were added. 60 MONOGRAPH IDRC-02le

TABLE 33. Comparative amino acid composition of TABLE 34. Percentage composition of all-mash chick triticale, maize, wheat and rye, calculated as amino starter diets (data from Bixler et al. 1968). acid/16 g N (data from Bixler et al. 1968). Triticale Maize Wheat Rye Amino acid Triticale Maize Wheat Rye Titricale, ground 72. 50 Ammonia 3.2 2. I 3.3 2.7 Maize, ground 62.21 Arginine 5.5 3.7 4.2 4.9 Wheat, ground 67.30 Histidine 2.2 2.5 2.3 2.3 Rye, ground 59.50 Lysine 3.1 2.6 2.5 3.8 Soybean meal Tyrosine I. 5 2.8 1.4 I. 5 (50~;, protein) 16. 15 27.44 21. 35 29. 50 Tryptophan I. 7 2.2 2.0 2.2 Other (similar in Phenylalanine 3.9 4.6 4.0 4.0 all diets) 11. 35 11. 35 11. 35 11. 35 Cystine I. 8 I. 7 I. 9 I. 7 Methionine 4.5 1.4 4.8 2.9 Serine 3.9 3.2 4.1 4.2 soybean protein (16.15%) in the triticale diet. At Threonine 3.1 2.9 3.0 3.7 that level, lysine would probably still be limiting in Leucine 6.0 12.2 6.2 5.9 the triticale diet. Isoleucine 3.2 3.4 3.2 3.3 Bragg and Sharby ( 1970) reported on three Valine 2.9 4.6 2.8 3.3 feeding trials to evaluate triticale, produced during Glutamic acid 27.3 17.9 30.6 23.7 1967, as a ~ource of energy and protein for broiler Aspartic acid 7. I 7.0 5.1 7.2 chicks, and to determine the utilization of amino Glycine 3.9 3.2 4.0 4.5 acids in triticale protein compared to wheat. Alanine 3.8 7.9 3.4 4.4 Each dietary treatment was arranged in a random Praline 8.9 8.3 9.7 8.9 block design and feed and water supplied ad libitum. Weight gain, F/G ratio and mortality In summary, on a weight for weight basis, the were calculated at 7-day intervals. The cereal triticale was approximately equal to hard spring portions of the basal, nutritionally balanced, wheat in nutritional value for chicks as judged by experimental rations, which were nearly isonitro­ weight gain, efficiency offeed utilization and ration genous and isocaloric, are given in Table 36. metabolizable energy. Lysine appeared as the In trial I, the three levels of triticale replace­ first limiting amino acid for chicks, with ment of wheat in the basal ration (A) (zero), (B) methionine being also limiting. 50% and (C) I 00%. were fed with three levels of Bixler et al. ( 1968) reported on their findings in dietary animal tallow, 0, 2.5 and 5% at the expense feeding trials using male chicks at Michigan of dextrose in the basal ration. Agricultural Experimental Station. The triticale, Although chick weight gain was slightly less grown in Sonora, Mexico was compared with from diets (B) and (C), the differences from the wheat and rye, also grown in Sonora and with a wheat diet (A) were not statistically significant. maize (de Kalb variety grown in USA) diet The addition of 2.5% animal tallow to the diets previously used at the Experimental Station. The amino acid composition of the grain (by auto­ TABLE 35. Nutritional value of triticale, maize, wheat analyzer) is given in Table 33. The percentage and rye in isonitrogenous diets for chicks (data from protein (Association of Official Agricultural Bixler et al. 1968). Chemists 1965), expressed on an as-received basis was triticale 16.75, wheat 12.94, rye 9.18. The Feed Feed diets were isonitrogenously balanced to give Weight consumption/ conversion approximately 23% protein. The cereal portions Grain gain, chicks, (feed of the four starter diets are given in Table 34. source 2 weeks 2 weeks consumed; The feeding results are summarized in Table 35. in the diet (g) (g) unit of grain) The maize and wheat diets were significantly Triticale 184.2 281. 2 I. 52 better than the triticale but no statistical dif­ Maize 208.3 279.3 I. 34 ference at 5% level of probability was found Wheat 208.0 296.8 1.43 between the rye and triticale diets. This result is Rye 174.8 269.4 I. 54 not surprising in view of the relatively low level of HULSE/LAING: PROTEINS OF TRITICALE 61

TABLE 36. Percentage composition of broiler chick trials, the protein content of 14.9% (N x 6.25) and diets (data from Bragg and Sharby 1970) the quality of the triticale, was very similar to that of hard wheat (protein 14.4%), which reduced the Diets influence of the higher protein triticale, previously (A) (8) (C) used, on the dietary amino acid pattern. ··--- The amino acid composition (by autoanalyzer; Wheat (protein (N x 6.25) 14.4':;,) 65.5 33.0 excluding tryptophan) of triticale and wheat and Triticale (protein (N x 6.25) 14.9° ) 33.0 66.0 0 the availability to chicks of the amino acids in the Soybean meal (protein N x 6.25) 44.0".,) 20.0 19.5 19.5 cereals (Bragg et al. 1969) are shown in Table 37. Other (similar in all diets) 14.5 14.5 14.5 Triticale and wheat protein were efficiently uti­

---·· lized with an average availability of 93.6% and 92.1 respectively. The methionine level of triticale (B) and (C), improved weight gain compared to was higher and utilized I 0% better than that of diet (A) but there was no significant difference wheat. The utilization of cystine was greater than between any of the three diets containing 2.5':., that of glycine in triticale but no significant fat. difference was observed in utilization among the The addition of 5':., fat to diet (B) significantly other amino acids. Glycine utilization in wheat improved the growth compared to diet (B) and was significantly less than the other amino acids. diet (C) without fat but was not significantly Methionine in wheat was utilized significantly

better than any diet containing 2.5~'o fat. The 5/0 better than glycine but was less available than the fat addition to diet (C) produced a severe growth other amino acids. depression due to a decrease of feed intake which Five triticale selections, grown at CIANO, was approximately 80% of the other rations. The identified as no. 24, 59, 106, 119 and 132, were F/G ratio was affected more by the addition of fat fed in chick feeding trials (McGinnis 1972). The to the ration than by the substitution of triticale triticales provided 8% of the protein in a 14% for wheat. There was no difference in metabolizable protein diet and were compared with a similar energy (Hill et al. 1960), of the three basal diets. diet in which 8% of the protein was provided by In trial 2, four supplemental levels ofoL-methio­ soybean meal. There were no significant differences nine (0, 0.05, 0.10 and 0.15%) were added to the between the triticales in chick PERs at two weeks. basal rations. Lysine was maintained at a constant No. 59, 119 and 132 were not significantly inferior level of I. I%· The addition of 0.05% methionine to the soybean meal diet though the PERs from to the triticale ration improved growth signifi­ No. 24 and 106 were significantly lower. cantly over the triticale control and the wheat The same triticale selections, fed in a practical control with no further improvement from the broiler diet in comparison with maize, gave similar higher levels of addition. No significant improve­ results to maize both when penicillin was, and was ment in F/G resulted from supplementation. not; added. In both the maize and triticale diets, In trial 3, four supplemental levels of DL­ the addition of penicillin resulted in significantly methionine (0, 0.05, 0.10 and 0.20%) and four improved growth. supplemental levels of L-lysine (0, 0.05, 0.10 and In another trial reported by McGinnis ( 1972) 0.20%) were added to the triticale basal diet (C), thirteen selections of triticales from CIANO were with and without methionine; a wheat control was compared, in chick diets, with soybean meal, included. Weight gain and F /G ratio were not Gaines wheat, "high protein" wheat (protein affected, and the significant improvement noted content not stated). and rye. The triticale selections in Trial 2 with 0.05% methionine was not repeated. were identified as no. 27, 37, 38, 39, 44, 48, 52, These differences are ascribed to the fact that two 53, 54, 61, 64, 65 and 67. The chicks on the soybean different strains of commercial chicks were used meal diet grew at a rate significantly higher than in the Trials 2 and 3. the chicks on any of the other diets. The triticale Sell et al. ( 1962) had found that chicks fed diets all gave higher rates of growth than those triticale rations responded positively to methionine obtained from the rye or from the high protein plus lysine treatment. Bragg and Sharby ( 1970) wheat, or from the Gaines wheat, with the excep­ state that the triticale used by Sell et al. ( 1962) had tion of no. 61, which was lower. Penicillin supple­ a protein content of 18.42% while in their own mentation improved significantly the growth 62 MONOGRAPH IDRC-02le

TABLE 37. Amino acid content and availability of triticale and wheat to growing chicks (data from Bragg and Sharby 1970).

Triticale Wheat

Content Availability* Content Availability* Amino acid (% total grain) (%) (% total grain) (%)

Lysine 0.439 93.4ab 0.362 94. 3c Histidine 0.296 9S.9ab 0.287 9S. Sc Arginine 0.710 92.0ab 0.604 92. Obc Aspartic Acid 0.879 92.4ab 0.687 91.9bc Threonine 0.421 91. 7ab 0.391 92.7bc Serine 0.621 94.7ab 0.609 94. Sc Glutamic acid 4.8Sl 97.3ab S.241 97.Sc Pro line 1 .196 97.2ab 1. 394 96.6c Glycine 0.673 8S.2a 0.627 70.8a Alanine 0.46S 90.3ab 0.394 89.9bc Cystine 0. 163 98. 3b 0.136 96. lc Valine O.S78 92.3ab O.S43 92. 2bc Methionine 0.204 90.0ab 0.180 81 .8b Jsoleucine 0.461 93.0ab 0.436 94. 2c Leucine 0.910 94.7ab 0.894 9S .2c Tyrosine 0.417 94.6a 0.384 94. 3c Phenylalanine 0.633 9S.8ab 0.64S 9S. 8c Tryptophan not determined Availability 93.6±4.66 92. l ± l. 90 (means of 17 amino acids) Protein (N x 6.2S) 14.9 14.4

*Means with the same letters are not significantly different (P < 0.01) within grain source. response to the rye diet but failed to produce 39% and 43% of a practical broiler diet, lysine statistically significant growth improvement from appeared to be the first and threonine the second, any of the other diets. The PERs and the feed limiting amino acid. efficiency paralleled the results for body weight. Trailblazer triticale was also used to provide McGinnis ( 1972) has also reported on chick 73% of the diet in broiler diets. Growth response feeding trials in which 8% of the protein in a 14 % was improved when the diet was supplemented protein diet was provided by soybean meal or by with lysine and when supplemented with penicillin. one of 27 triticale selections provided by A significantly greater response was achieved Washington State University, the remaining 6/o when both were used together, the diet being then protein being provided by a premix. A control equal to the control diet containing maize supple­ diet, in which all the protein was provided by the mented with penicillin. premix, was also i11cluded. The soybean meal diet Turkey Pou/ts and Laying Hens Feeding Trials gave better growth than any of the triticale diets. Sell and Johnson ( 1969) examined triticales from The triticales together with their protein contents two suppliers in North Dakota as feed for turkey (Kjeldahl N x 6.25) are listed in Table 38. The poults and laying hens. Triticale A had been selections which were most effective in supporting cleaned and was known to be relatively free of chick growth are indicated by an asterisk. ergot; triticale B was in crushed form and con­ Trailblazer triticale of protein 18.6% (N x 6.25) tained a considerable quantity of weed seed; was also included in the trials. When supplemented possible contamination with ergot could not be with amino acids to a level equal to the soybean determined. meal diet, the triticale-sustained chick growth was In the turkey poult trials, both triticales were not significantly lower than growth on the soybean compared with a durum wheat on a weight for diet. When Trailblazer was used at levels between weight basis in nutritionally balanced turkey HULSE/LAING: PROTEINS OF TRITICALE 63

TABLE 38. Protein contents of triticale selections from wheat and triticale A were similar, that of triticale Washington State University (data from McGinnis B was significantly lower. 1972). Sell and Johnson conclude that clean triticale, relatively free of ergot, compares favourably with Triticale Protein (N x 6.25) wheat in nutritional value for poultry. selection e;,i Guenthner and Carlson ( 1970), compared triti­ RS45 15.8 cale, maize, wheat and sorghum laying diets (the LNTC-1 19.0 exact composition is not given) when fed to single RS46 15.7 comb White Leghorn hens. Two diets were formu­ 6TA487 16.6 lated from each grain to contain 12.0% and 15.4% *RS47 14.4 crude protein (N x 6.25). All the protein in the *6TA484 15.2 12% wheat and triticale, essentially isocaloric, *RS48 13.2 diets was provided by the cereals, the other diets *6TA476 16.2 *RS49 14.5 being balanced with soybean meal. Methionine 6TA190 16.2 and lysine were added to provide a minimum of 6TA199 17.8 0.52% methionine plus cystine and 0.50% lysine. 6TA494 17.4 Egg production performance was judged on the 6TA385 17.5 averages of three weeks preceding the nine week 6TA327 18.3 test period and the final three weeks. Initial CTC-4 17. 1 average egg production for all groups at 34 weeks *6TA518 17.3 of age was 75%. In the 15.4% protein series egg *6TA131 18.8 production increased 4% with wheat and maize, *6TA480B 16.8 5% with sorghum and decreased 4% with triticale. *6TA492 17.0 6TA491 16.8 In the 12% protein series, production decreased *6TA480A 16.2 13~/0 with triticale, I 0% with wheat, 7% with *6TA204 16.7 sorghum and increased 4% with maize. Hen per­ 6TA418 16.7 formance on both maize diets was equal; the 12% 6TA488 16.5 protein triticale, wheat and sorghum diets did not *6TA479 16.6 support egg production as well as did maize. Hens 6TA205 16.3 on both the triticale diets and on the 12% wheat *6TA486 16.3 diet lost weight, indicating amino acid deficiencies other than lysine and methionine plus cystine. The *Most effective in supporting chick growth. effect of the superior amino acid composition of the protein in the soybean meal used to balance starter rations, the grains representing 42% of the the maize diet possibly influenced this result. test diet. The weight gain of poults on all three rations from one day to three weeks of age was Weber and Reid ( 1971) compared two un­ approximately equal indicating little, if any, ergot specified types of triticale and four varieties of contamination of the triticale. Efficiency of feed wheat with sorghum in laying hen diets consisting utilization (F/G) was similar for poults fed durum of 17% crude protein (N x 6.25) and metabolizable wheat and triticale A but was significantly less energy values of 2780 kcal/kg. Differences in efficient with triticale B. The metabolizable energy egg production and egg weight, but not in shell (ME) contents of the wheat and triticale A were thickness, were found but no further details are similar but that of triticale B was relatively low; given. more feed being required per unit of weight gain. Two triticales, no. 204 and a composite of no. The trial was continued with poults from three 418 and 419, were compared with the wheat to six weeks of age but only durum wheat and varieties Sonora 64, Siete Cerros and INIA 66. triticale B could be compared as there was and two sorghums over 252 days in feeding trials insufficient triticale A. Again weight gains were with laying hens at the University of Arizona essentially similar but the F/G ratio was less (Weber et al. 1972). Siete Cerros and the triticale efficient from triticale B. composite gave results in egg production and feed The same grains were included at a level of 53% conversion similar to sorghum; Sonora 64. INIA in rations for laying hens. The ME of durum 66, and Triticale 204 all lowered egg production 64 MONOGRAPH IDRC-02le

significantly. All the wheat and triticale samples Triticale, of approximately 15% protein, of the produced lower egg weights than the sorghums. 1962 and 1964 crop years was fed to Managra When, in a second experiment, !NIA 66, Siete gilts and barrows and five Yorkshire barrows in Cerros, Triticale 203, Triticale 204 and a com­ finishing rations. The amino acid content of all mercial sorghum were compared, egg production, the triticale used was, except for glycine, similar feed conversion and egg weights were similar. to that of hard spring wheat but there was in the Metabolizable energy values averaged 3130 1964 triticale crop a 30% decrease in methionine kcal/kg for the triticales and 3450 kcal/kg for the compared to the 1961 crop. wheats. Stothers and Shebeski report a marked palat­ A diet for laying hens in which triticale of ability problem with the 1962 triticale. When 1962 unstated origin provided 82% of the protein with crop triticale formed 50% of the cereal portion of the remainder of the protein coming from fish­ the finishing ration, average daily feed con­ meal and alfalfa gave results equivalent to those sumption for both gilts and barrows was approxi­ from a conventional layer diet. However, when mately 5 lb (2.2 kg) per pig per day. Feed con­ fishmeal was not included and the protein was sumption for the first nine days of the finisher test supplied by triticale, alfalfa and wheat, and was approximately half this level. When the feeding supplemented with lysine and methionine at levels level of the barley ration was restricted to 5 lb calculated to be adequate, egg production was (2.2 kg) per pig per day, average daily gain, feed poorer than from the control diet (Fernandez et al. efficiences and carcass data approximated to those 1972). of the pigs fed the triticale ration, suggesting the Swine Feeding Trials Stothers and Shebeski presence of an appetite depressant within the (1965) conducted trials with hexaploid triticale triticale ration. grown in the crop years of 1961, 1962 and 1964 When gilts were fed triticale as a sole finishing as a feed for growing swine. grain, average daily feed consumption dropped to The protein content (analytical method not 0.8 lb (0.3 kg) for the first nine days. Reduction stated) of the 1961 crop was 19% and three starter of the triticale level to 20% for eight days improved rations on an isonitrogenous basis (approximately feed consumption to 4. 7 lb (2.1 kg), an increase 19% protein) were formulated in which (A) barley, to 50% level for 21 days further improved feed (B) barley 50%, triticale 50% and (C) all-triticale, consumption to 5.6 lb (2.5 kg) but a subsequent made up the cereal grain portion. Gilts and increase to I 00% triticale reduced feed con­ barrows of the Managra strain were used; in the sumption to 4.4 lb (2.0 kg). Barrows were fed first part of the test initial weights were approxi­ rations containing gradually increased levels of mately 35 lb (16 kg), and in the second part, initial triticale, 20% for two days, 50% for eight days and weights were approximately 86 lb (39 kg). Water 100% for an average of 54 days. Average daily and feed were given ad libitum. feed consumption was 2.5, 4.5 and 4.5 lb (I. I, 2.0, The results indicated that the triticale was used 2.0 kg) respectively. The final shift to I 00% satisfactorily by pigs started at the heavier weights triticale resulted in both barrows and gilts showing but not by those started at the lighter weights, marked lack of appetite which did not recover to when fed over a 4-week period. full feeding level. In early 1962 grower tests were conducted with The 1964 crop gave essentially similar results. Managra barrows and gilts fed from initial weights Two lots of barrows and gilts were fed either a of 68 lb (30.8 kg) to final weights of 125 lb (56.8 kg). barley finisher ration or a triticale ration, the lots The grower rations again contained (A) barley, being subsequently reversed. With both barrows (B) barley 50%, triticale 50% and (C) all-triticale and gilts, feed consumption dropped to almost as the cereal grain portion. A finisher test was con­ nil when triticale was the sole cereal grain but a ducted comparing all-triticale with all-barley ration which contained 30% triticale and 70% finisher rations using barrows initially weighing barley was satisfactory. Pelleting the triticale had 132 lb (60 kg). Insufficient supplies of triticale no apparent effect. limited this particular test. Shimada et al. (1971) compared hexaploid triti­ The growing and finishing tests for 1961 crop cale and sorghum in rations for growing swine in showed no significant differences in average daily Mexico. The triticale used was a mixture of several gain among rations. No carcass abnormalities varieties produced in 1970 with a crude protein were apparent in the animals fed triticale. content of 16.1 % dry basis equivalent to 14.1 % HULSE/LAING: PROTEINS OF TRITICALE 65

TABLE 39. Essential amino acids of triticale (as kg received the same basal diet as in the second percenr of 1rhole drr grain) and of a typical sorghum trial with and without 0.16% added L-lysine and used in feeding trials with growing swine (data from 0.10% DL-methionine in a 2 x 2 factorial arrange­ Shimada et al 1971 ). ment of treatment in four completely randomized ------replications. The addition of lysine resulted in a Sorghum highly significant improvement in average daily (Data from National gains and F/G. Addition of methionine alone Amino acid Triticale Research Council 1969) produced a slight depression in weight gain and Arginine 0.97 0. 38 F/G which was not significant. When both amino Histidine 0.38 0.26 acids were added weight gain increased but the lsoleucine 0.62 0.49 combined effect was not statistically significantly Leucine I. 21 1.60 better than the addition of lysine alone. Lysine 0.57 0.25 Different results were obtained for the common Methionine 0.09 0.15 treatment in the two trials. The triticale used and Phenylalanine 0.80 0.60 the initial body weight of the pigs were the same Threonine 0.55 0. 37 but the breed of pigs, Duroc in the second trial and 0.16 0.10 Tryptophan Hampshire in the third trial, and the time of year, Valine 0.82 0.60 fall vs. winter, were different. In summary, triticale was substituted suc­ cessfully for sorghum and for part of the soybean protein as fed at a moisture content of 12.5%. The meal in diets for growing pigs. sorghum had a crude protein content of 10.9% Harrold et al. (1971) evaluated hexaploid dry basis, 9.6% as fed at 12.0% moisture. The triticale from two crop years as a feed for growing­ amino acid content (by autoanalyzer) of the finishing swine. The basal rations included soy­ triticale and of a typical sorghum is given in bean meal 10.9% and (A) barley 86.9%, (B) barley

• Table 39. 43.5%, triticale 43.5%, and (C) triticale 86.9/0 In the first trial, 16 Duroc gilts with an average All the rations were ground and pelleted. The initial weight of 14.6 kg received a diet in which crude protein content of the rations was (A) 15.5%, triticale was progressively substituted for the (B) 16.3% and (C) 16.7%. grain in a 16% protein sorghum-soybean meal diet. In the first trial, the three basal rations were fed The diets were made isonitrogenous by adjusting to six pigs having an average initial weight of the content of soybean meal. Substitution of about 56 lb (25.4 kg). The triticale, grown in triticale at 30, 60 and 90% for the sorghum did not 1968, had not been cleaned and contained ergot. significantly affect the performance. Shimada et al. The experiment ended after 91 days. Inclusion of (1971) cite Bowland (1968) as reporting a similar the triticale produced a marked reduction in feed result when triticale replaced 50 and 100% of the intake and average daily gain, the most severe wheat and barley constituents in diets for growing effects resulting when the triticale was the sole pigs. grain source. In the second trial, 12 Duroc pigs, six gilts and In the second trial, the cereal portion of the six boars, average weight 12.2 kg received a diet rations were (D) barley, (E) barley 50% and in which L-lysine HCI (0, 0.08 and 0.16% of triticale 50% grown in 1969, (F) triticale grown in L-lysine) was added to a 96% triticale basal diet. 1969, uncleaned, (G) triticale grown in 1968, There was a significant linear improvement in cleaned. Rations were fed to two lots of six pigs weight gain with the addition of lysine. The feed: with an average initial weight of about 66 lb gain (F/G) ratio also improved but the relation (30.0 kg). The experiment ended after 45 days. was not statistically significant. The total calcu­ The 1969 triticale produced results comparable lated lysine content of the diet with the highest to those obtained from the barley ration. Cleaned addition of L-lysine HCI was 0.65% which is still triticale from the 1968 crop was inferior to barley less than the optimum 0.7% suggested (Germann and triticale from the 1969 crop as the sole grain et al. 1958) as being required for 11.4 kg pigs fed for growing pigs. The F /G ratios were not markedly 13.4% protein diets. different. In the third trial, 16 Hampshire pigs, eight In the third trial, the cereal portion of the rations gilts and eight boars, average initial weight 12.3 were as in the second trial except that the triticale 66 MONOGRAPH IDRC-02le

TABLE 40. True nitrogen digestibility of soybean, general triticale appeared to be somewhat un­ triticale, wheat and barley for I 0-kg and 30-kg pigs palatable to growing-finishing swine. (data from Sauer 1972). Gundel et al. (1970) compared two hexaploid triticales, designated 57 /69 and 64/69 grown in True nitrogen digestibility. 0 ~ Hungary, in diets for pigs of weights varying from 40 to 60 kg and concluded that triticale could be IO-kg pigs 30-kg pigs used successfully in their feed. Soybean 94.7 91.6 Sauer (1972) compared biological availability Triticale 92.0 89.6 (de Muelenaere et al. 1967) to Managra barrows. Wheat 90.9 88. l weighing I 0 kg or 30 kg at the start of the trials, of Barley 85.8 88.7 16 amino acids (tryptophan and cystine not included) in triticale, wheat, barley and soybean, all of unstated origin. The faecal analysis method was uncleaned from the 1969 crop. Each ration of Kuiken and Lyman (1948) was employed; was fed to three pens of six pigs with an average amino acid analysis by the method of Bragg et al. initial weight of about 86 lb (39.0 kg). It ended (1966). after 77 days when the average weight of the The true nitrogen digestibilities are shown in heaviest group exceeded 200 lb (90.0 kg). Average Table 40 and the mean true essential amino acid daily gain and feed intake was again highest for availabilities in Table 41. Among the amino the pigs fed the barley ration and lowest for those acids, lysine was the least available in the cereals. fed the ration with all triticale as the cereal source. Among the cereals, the essential amino acids were In summary, Harrold et al. recommended that most available from triticale, followed by wheat, triticale be limited to a maximum of 25% of thP followed by barley. ration for growing pigs (50 to 130 lb; 22.8 to 59 kg) Cornejo et al. (1972) at Davis, California, or 50% of the grain mixture for finishing pigs ( 130 compared a triticale of 15.3% crude protein lb (59 kg) to market weight). It is not recom­ (N x 6.25) with commercial wheat, barley and mended for breeding stock or young pigs. In maize in feeding growing-finishing swine. The

TABLE 41. Mean true essential amino acid availabilities (excluding cystine and tryptophan) of soybean, triticale, wheat and barley for IO-kg and 30-kg pigs (data from Sauer 1972).

Mean true availability,%

Soybean Triticale Wheat Barley

IO-kg 30-kg IO-kg 30-kg 10-kg 30-kg IO-kg 30-kg pigs pigs pigs pigs pigs pigs pigs pigs

Least available in cereals Lysine 94.9 91. 2 86.3 77. 5 80.8 67.3 77. l 65.0 Intermediate availability in cereals Isoleucine 95.2 92. l 91. 5 87.7 89.2 86. l 84.7 83. I Methionine 94.5 86.3 90.3 86.8 89.8 85.7 87.3 81. 5 Threonine 95.4 91.0 90.9 87.7 87.3 85.4 87.5 82.5 Valine 94.5 90.9 92 .4 86.8 89.0 85.9 87.6 84.6 Tyrosine 95.0 93. l 92.9 89.7 90.3 87.5 87.6 85.0 Leucine 95. 7 93.2 93.7 91. 3 91. 2 89.8 88.7 87.6

Most available in cereals Arginine 98.4 96.0 94.5 94. l 94. l 93. I 91. 4 90. l Histidine 97.9 94.6 93.8 93.3 94.7 92.7 93.2 89.5 Phenylalanine 95.8 94.2 94.5 93.3 92.4 91.6 90. l 89.5 HULSE/LAING: PROTEINS OF TRITICALE 67

), TABLE 42. Digestible energy (DE), metabolizable energy (ME), corrected ME (ME 0 and nitrogen (N) reten­ tion values of triticale, wheat, barley and maize for growing-finishing swine (data from Cornejo et al. 1972).

N retention DE (kcal/kg o,

dry matter) ME ME 0 ingested N absorbed N

Triticale 3,603 3,522 3,225 34.4 40.2 Wheat 3,709 3,625 3.335 33.2 37.9 Barley 3,375 3,332 3,052 39.9 51. 9 Maize 3,796 3,745 3,560 33.6 41. 5 diets were composed of 95% cereal. The experi­ amino acid in triticale for finishing swine, and also ment was conducted to determine digestible suggest that the lysine in triticale may not be fully energy (DE), metabolizable energy (ME) and available to the pig.

) corrected ME value (ME0 as well as the ability Allee and Hines (l 972b) also compared triticale of triticale to promote nitrogen retention. The of 15.96% protein (conversion factor not stated) values obtained, presented in Table 42, indicate and 0.625% lysine on a moisture-free basis, in that triticale is comparable to wheat and maize meal and pellet form in diets for growing pigs and superior to barley as an energy source, and weighing 26 kg. The diets were: (A) (control), able to support a high positive nitrogen balance. sorghum and soybean meal, 16.5% protein, (B) Chubb (1972 unpublished data) compared triticale replacing 20% of the sorghum, (C) 40% triticale of nitrogen content 2.06 with ground replacement, (D) 60% replacement, (E) 80% barley, and naked or hull-less barley, variety replacement, (F) 100% replacement, all in meal Nackta, in isonitrogenous diets for pigs. The basal form, (G) 100% replacement in pellet form, diet included, in percent, barley meal 71.0, (H) triticale and soybean meal, isonitrogenous soybean meal extract 7.5, white fishmeal 2.0, with diet (A), (I) triticale + 0.2% L-lysine. glucose monohydrate 16.0, minerals, etc., 3.5. In The level oftriticale and the physical form, meal the experimental diets, the test cereals were or pellet, had no significant effect on feed intake, substituted for 25% of the basal diet. The apparent daily gain or F/G ratio but pigs fed diet (H) (iso­ digestibility of dry matter and nitrogen and the nitrogenous with diet A) and diet (I) gained gross energy of the triticale and Nackta diets were significantly more slowly than those on the other greater than those of the barley diets. diets. The results indicate that triticale may be Allee and Hines (l 972a) fed triticale of 18.41 % substituted for sorghum on a weight for weight protein (conversion factor not stated) and 0.625% basis, but not on an isonitrogenous basis, in lysine, both expressed on a moisture free basis, in diets for growing pigs. finisher rations to pigs weighing 49 kg. The diets, These findings appear to disagree with those of all in pellet form, were: (A) triticale alone, (B) Shimada et al. (1971 ). triticale + 0.1 % lysine, (C) triticale and soybean meal, (D) sorghum and soybean meal, (E) wheat Cattle and Sheep Feeding Ingalls et al. and soybean meal. Diets (A), (D) and (E) contained (1970) compared the feed in take and growth rate the same level of lysine and all diets received the of young dairy calves receiving two levels of same mineral, vitamin and antibiotic additions. triticale (27.5 and 55% of the complete ration) There were no significant differences in daily gain, with calves receiving barley diets supplemented feed intake or F/G ratio among the pigs fed diets with urea or soybean meal. The triticale was from (B), (C), (D) and (E) but pigs fed diet (A), triticale the 1967 crop and the ergot content of the 55% alone, gained weight significantly more slowly triticale diets ranged from 0.03 to 0.11 % and was than pigs on the other diets. Carcass analysis proportionately lower in the 27.5% triticale diet. showed no significant differences among diets. The highest level of 0.11 % just reached the level The results indicate that lysine is the first limiting which might be considered dangerous. The grain 68 MONOGRAPH IDRC-02le

was fed dry rolled but a further ration with 55/0 cass traits of finishing steers, 40 Hereford steers, pelleted triticale was also fed. average weight 285 kg, were randomly allotted The rations were not isonitrogenous, the percent into two main treatment groups with five replica­ crude protein contents on a dry matter basis being tions of four steers in each treatment. The rations

(A) barley-urea 16.8%, (B) barley-soy 16. 7/0 , were isonitrogenous at 15.1 % crude protein as fed,

(C) triticale (27.5% level) 18%, (D) triticale (55/0 and contained 7% roughage. Dry rolled triticale level dry rolled) 18.5%, (E) triticale (55% level comprised 92% of that ration. The steers were pelleted) 18.9%. self fed, were weighed at 28-day intervals and Ten dairy calves, five bulls and five heifers were slaughtered on completion of the 146-day feeding assigned to each of the experimental diets which period. The average daily gains were significantly were fed once daily in amounts such that feed was higher among steers fed the sorghum ration than available at all times from one week of age. Water among those fed triticale and there was a lower was supplied ad libitum. The calves were weighed consumption of triticale. Feed conversion of the at 2-weekly intervals and removed from the triticale ration was more efficient. Standard carcass experiment when they weighed more than 137 kg. traits were similar, but more condemnations Daily dry matter intake from six weeks of age because of liver abscesses occurred among steers to final weight was 9 to 11 % greater for diets (A) fed triticale. and (B) than for diets (C) and (D) for which intake In a trial to compare digestibility, 12 steers of reduction was similar. Pelleting the 55% triticale average weight 272 kg and 14 crossbred wethers ration, diet (E), resulted in a further 12.6% reduc­ averaging 43 kg were randomly allotted six and tion. seven per treatment respectively to rations con­ Daily weight gains for the same period were 12 taining dry rolled triticale (79%) or sorghum to 16% less for calves on diets (D) and (E) than for (71.6%) and roughage. A 7-day adjustment period those on diets (A) and (B). There was no difference was followed by 7-day preliminary and collection in weight gain between calves receiving diets (C) periods. AH~nimals were initially fed the experi­ and (D). Pelleting the triticale. diet (E). did not mental rations on a limited daily intake basis affect weight gain, and feed efficiency was therefore (1% of body weight) in two feedings and intake greater for the pelleted triticale than for the dry gradually increased during the adjustment period. rolled, and was not significantly different from Maximum consumption was limited at 6.35 and diets (A) and (B). Differences among diets for 1.20 kg per head daily for the steers and wethers apparent dry matter and protein digestibilities respectively. Feed intake was held constant at the were not significant. Bull calves demonstrated level established for each animal during the greater gains, feed intake and efficiency than did remainder of the trial. Water was provided ad heifer calves. libitum. Both apparent and true digestibility of In summary, complete calf rations containing crude protein were significantly higher for the 27.5 or 55% dry rolled triticale resulted in a small triticale ration. reduction in feed intake and weight gain com­ It had been observed early in the finishing trial pared with barley-urea and barley-soy rations but that consumption was lower with the triticale feed efficiency was similar. Pelleting the 55/0 ration, and it had also been found that intake triticale ration resulted in lower intake, equal levels in the digestibility trial were more difficult weight gain and increased feed efficiency com­ to establish with triticale, especially with sheep. pared to the dry rolled triticale. Low levels of Therefore three 28-day acceptability trials, with ergot contamination of the triticale may have four treatments in each trial were carried out using affected feed consumption. 16 Hereford heifers initially averaging 227 kg. McCloy et al. (1971) compared a mixture of There was a 3-week rest period between trials and hexaploid triticales, crude protein 17.8% dry no heifer received rations with identical triticale matter basis, grown under irrigation in California levels during any two successive trials. The heifers and Texas in 1969, with commercial sorghum, were fed individually in all three trials. Average crude protein I 0.9% dry matter basis (N x 6.25). daily consumption expressed as kilograms per The triticale was free from detectable ergot. mean metabolic size, determined from an average In a trial to compare performance, feed con­ of initial and final weights, was used to evaluate sumption, efficiency of feed utilization, and car- comparative acceptability of the rations in each HULSE/LAING: PROTEINS OF TRITICALE 69 trial. In the first acceptability trial the rations Crude protein digestibility was lower with Sturdy contained 0, 30, 60 and 90% dry rolled triticale wheat than with the other diets. Nitrogen retained replacing equal parts by weight of sorghum and as percentage of intake tended to be lower with the other concentrates. All rations were isonitrogenous Winter triticales than with the Spring triticales and and contained 9% roughage. Consumption de­ the wheats, but overall the results indicated that creased at each higher level of triticale, with a Spring and Winter triticales had comparable significant decrease between the 30 and 60/0 levels. nutritive value for sheep with Sturdy wheat lower In the second acceptability trial rations were and Tascosa wheat higher than the triticale. similar except that molasses replaced 5/0 by Triticale grown in the Texas Panhandle area weight of the grain; in rations containing both during 1970 was compared with barley grown in grains, each grain was reduced in proportion to Arizona on an equal weight basis in feeding trials its level in the ration. There was also a significant with lactating (60 to 120 days post partum) decrease in consumption when the rations con­ Holstein cows (Moody 1973). The rations were tained 5% molasses. fed individually, and consisted of cubed alfalfa at In the third acceptability trial dry rolled 1.8% of body weight daily, with the balance of the triticale and sorghum were compared in rations net energy requirements being provided by con­ with I 0% roughage and constant grain: roughage centrates. The concentrates consisted of 92% ratios, with and without molasses. Consumption grain, 7% molasses and I% salt. The protein values was significantly lower with the triticale rations. (conversion factors not stated) of the rations as The addition of molasses did not influence con­ eaten were: (A) steam rolled triticale 18%. (8) sumption within grain treatments. steam rolled barley 15.5%. (C) equal amounts of McCloy et al. (1971) cite Lofgreen (1969) who triticale and barley 16.9%. All rations were found that feed intake and gains were higher and equally well accepted and no significant differences feed utilization more efficient with triticale as were found, attributable to the grain source, in compared with sorghum, when both were steam milk yield and composition, ruminal volatile fatty rolled and fed in rations containing 68% grain and acids, or digestibility. 7% molasses. The triticale was from the same Rosner triticale silage was evaluated against source as that used by McCloy et al. ( 1971 ). maize silage in the diet of lactating Holstein cows McElroy (1968), also cited by McCloy et al. (Fisher 1972). On the triticale silage diet, a marked (1971 ), found that steers fed triticale displayed depression in milk protein content, together with higher and more efficient gains than did steers fed lower milk production and loss in body weight was barley, and that consumption of steam rolled reported. The apparent digestibility of dry matter triticale rations was higher than dry rolled triticale. for rations containing triticale silage was 64% He also reported a higher incidence of liver and for maize silage 67%. abscesses when triticale made up more than 50~, Gundel et al. ( 1970) compared two hexaploid of the grain portion in finishing rations, and con­ triticales, designated 20/68 and 57 /68 grown in siderable damage to ruminal epithelium in steers Hungary, in diets for wethers and concluded that fed triticale at high levels. triticale could be used successfully in sheep diets. Sherrod ( 1972) compared two winter triticales designated 131 and 385, two spring triticales, 204 and 208, and two hard red winter wheats, Sturdy Environmental and Agronomic Effects and Tascosa, in 50% grain rations, fed at 800 g daily, dry rolled with chopped forage sorghum The influence of environment and fertilizer hay, to sheep in digestibility studies. Crude treatment on the protein quality of triticale is protein levels, dry matter basis, were comparable evidenced by data provided by Dr. Eva Villegas in among the triticales at 22% and between the wheats June 1970, and presented by Zillinsky and Borlaug at 18%. All rations were eaten readily. Digestibility (197la). The protein and lysine content of of the major non-fibre energy components was not identical strains from 11 crosses grown at CIANO significantly different among rations; crude fibre in 1968-69 and at Navojoa in 1969-70 are com­ digestibility was comparable among the triticales pared in Table 43. The average protein content was with Sturdy wheat significantly lower and Tascosa higher and the percent lysine in the protein lower in wheat significantly higher, than the triticales. the seed from the CIANO crop. Test weight and 70 MONOGRAPH IDRC-021e

TABLE 43. Protein and lysine content of triticale crosses grown at two locations under different levels of nitrogen fertilizer (data from Zillinsky and Borlaug 1971 a).

Average~~ Average% lysine Lysine in protein in protein sample Cross No. of Number strains Y68-69• N69-70b Y68-69• N69-70b Y68-69• N69-70b

x 136 3 16.85 14.84 3.00 3.70 0.505 0.549 x 160 16.30 15.39 3.25 3. 12 0.529 0.480 x 195 5 16.34 15.65 3. 16 3.18 0.516 0.497 x 224 13 17.78 16.03 2.67 3.22 0.475 0.516 x 281 3 16.59 16.62 2.72 3.03 0.451 0.504 x 284 6 16.92 14.81 2.56 3.34 0.433 0.495 x 298 6 16.33 15.27 2.71 3.11 0.442 0.475 x 308 57 15.63 14.35 2.94 3.20 0.459 0.459 x 313 18 16.30 14.17 3.28 3.35 0.534 0.475 x 653 37 16.44 15 .19 2.77 3.29 0.455 0.499 x 674 2 16.58 15.62 2.80 3.30 0.464 0.515 Average all samples 151 16.26 14.96 2.83 3.25 0.460 0.486

•cJANO winter nursery 1968-69; 120 kg/ha/N application. bNavojoa winter nursery 1969-70; 60 kg/ha/N application. seed grains were similar for the two seasons but increased the yield significantly over the control. lodging was more serious on the CIANO crop. The protein content of about 20% dry weight All the varieties oftriticale used by Villegas et al. (conversion factor not stated) and 1000-kernel ( 1970) (see Table 18), were grown at one location, weights, were unchanged by all treatments. and the variations should therefore reflect varietal differences. Szabo ( 1972) reported that hexaploid triticale The first section of this chapter has dealt with had been successfully grown in Hungary, triticale and has included papers in which triticale especially in sandy areas. From experiments with was compared with wheat and/or rye. five strains of Hungarian grown triticale to ascer­ The subsequent sections of the chapter deal tain germination requirements, he reported that with (a) wheat alone, (b) comparisons of wheat triticale grains are more sensitive to higher con­ and rye and (c) rye alone. centrations (2 to 3%) of salt solutions than simultaneously germinated rye grains. The salt tolerance of triticale is roughly equivalent to that Wheat of wheat and oats. The optimum germination temperature of the varieties examined was 20°C. Genetic Effects the maximum being 35°C (40°C in the case of one variety) and the minimum 5°C. TOT AL PROTEIN Tennenhouse and Lacroix ( 1972) have reported The influence of nitrogen metabolism and trans­ on the effect of treatment with (2-chloroethyl) location of nitrogen in the wheat plant upon seed trimethylammonium chloride (CCC) at 3.0 kg/ha protein content has been reported upon by and 6.0 kg/ha with and without nitrogen at 90 Johnson et al. ( 1967), and Johnson and Mattern kg/ha on Rosner triticale. Both levels reduced the ( 1972). Nitrogen metabolism and the nitrate height of the mature plant, the higher level produc­ reductase system has been discussed by Croy and ing greater reduction, but the reduction was less Hageman ( 1970) and Duffield et al. ( 1972). when nitrogen was added. There was no lodging. Field experiments to estimate the broad-sense The 6.0 kg/ha level of CCC on the unfertilized plot heritabilities in wheat, on an individual plant HULSE/LAING: PROTEINS OF TRITICALE 71 basis, conducted at Oklahoma State University, TABLE 44. Mean, standard deviation and heritability showed that nitrate reductase activity was sig­ of grain protein, flour yield and flour protein (data nificantly correlated with grain protein, water­ from Baker et al. 1971 ). soluble protein, and nitrate content. NB 65679, a selection from a cross between Atlas 66 and Wheat Comanche, obtained from the Nebraska Agri­ grain Flour Flour cultural Experiment Station, was a high protein protein yield protein wheat used in these trials. (%) (%) (%) The results indicated that higher nitrate reduc­ Mean. average of five tion and more efficient nitrogen translocation estimates 14.56 75.6 13.80 were involved in producing higher protein in the Standard deviation, average grain. The reductase system appeared sensitive to of four estimates 0.20 0. 56 0.18 both environmental and physiological influences Heritability (Duffield et al. 1972). (estimates significantly greater Rao and Croy ( 1972) studied the protease and than zero (P = 0.05) if they nitrate reductase patterns in three .. high protein" exceed 0.46, 0.48, 0.50 and 0.44 wheats, each having Atlas 66 germ plasm in their in each of the four sets of data, respectively) parentage, in comparison with Triumph 64, a 1965-66 0.86 0.81 0.92 hard red winter "low protein" wheat widely grown 1966-67 0.84 0.58 0.87 in Oklahoma. There were no significant differences 1967-68 0.61 0.46 0.76 in nitrate reductase and protease activity levels 1968-69 0.89 0.80 0.95 among the three high grain protein wheats but Average 0.80 0.66 0.88 the high grain protein wheat variety NB 65317 (selected as representative) showed higher nitrate reductase activity prior to heading, and higher location interaction peculiar to each year. The protease activity after heading, than did the lower estimates indicate that protein content is highly grain protein wheat Triumph 64. heritable but Baker et al. state that the heritability Baker et al. ( 1971) analyzed quality data for estimates are biased upward due to their inability hard red spring wheat cultivars grown in Canada in to account for genotype x location interactions. Western Cooperative Wheat Tests in the five years There was a high positive correlation (0.96) 1965 through 1969. In each year from 23 to 25 between flour and grain protein indicating that cultivars were grown at 15 to 16 locations. Among milling had essentially no differential effect on the quality traits examined for heritability were flour protein in the cultivars studied. grain protein and flour protein (American Associa­ tion of Cereal Chemists 1962). The heritability was AMINO ACID COMPOSITION estimated from analyses of variance of data from In the view of Mertz ( 1971 ), the chances of in­ (A) the 13 cultivars that were common to 1965 creasing the quantity of protein in wheat and and 1966, (B) the 12 cultivars common to 1966 triticale appear to be greater than the chances of and 1967, (C) the 11 cultivars common to 1967 increasing the biological quality of that protein. and 1968, (D) the 14 cultivars common to 1968 However, encouraging results are emerging from and 1969. Grain was milled, straight grade, in an the research being conducted at the University of Allis-Chalmers mill. Heritability was estimated as Nebraska into the nutritive quality of wheat the ratio G/(G +£),where G was the component through increased protein content combined with of variance due to average (genetic) differences improved amino acid balance. Since lysine is the among cultivars and E was the component due essential amino acid in shortest supply in cereals, to deviations from average performance (measure­ most attention is being given to increasing the ment errors and genotype x year interaction). proportion of lysine while maintaining a high Means, standard deviations and heritability esti­ grain protein content. mates are presented in Table 44. The standard The studies at the University of Nebraska have deviations reported include the error of the been conducted over many years, and are reported laboratory analysis and the effect of genotype by in several publications: Haunold et al. ( l 962a, b); 72 MONOGRAPH IDRC-02le

Johnson et al. (196 7, I 968a, b, 1969, 1970, 1972); unit dry weight of the grain, the high protein Mattern et al. ( 1968, 1970); Johnson and Mattern group provided more of every essential amino (1972). acid than the high lysine group. Among the objectives of the Nebraska research Potentially useful genetic sources of high pro­ are (I) to analyze systematically the wheats, both tein and/or high lysine are shown in Table 45. T. ru/gare and T. durum, in the world wheat Several of the protein sources may carry the same collection for protein and lysine contents, and (2) genes for protein. No gene for high lysine com­ to incorporate genes for high protein and high parable to the opaque-2 mutation in maize has lysine into wheats potentially suitable for cultiva­ yet been identified in wheat but the sources given tion in developing countries. It is a further purpose in Table 45 have been consistently superior in to distribute potentially superior germ plasm to lysine to other varieties examined. In greenhouse interested developing countries for adaptability tests, Nap Hal (Pl 176217) has proved superior and breeding research. in both protein and lysine contents (Johnson and The results of the systematic screening of the Mattern 1972). world wheat collection were reported as they became available (some 47 publications have been issued between 1966 and 1972) and have been Varietal Influence summarized by Johnson and Mattern ( 1972). More than 12,600 common wheats (T. ru/gare) and TOT AL PROTEIN 3,400 durum wheats ( T. durum) have been An inverse ratio between grain yield and protein analyzed for protein content and amino acid content was noted by Williams (1966) who composition with special attention to lysine. reported that in a series of 24 trials from 1961 to Because of the large number of samples of 1963, throughout the Australian wheat belt, the relatively small size to be examined, special wheat variety Festival was consistently high in analytical techniques have had to be developed. protein and low in yield while Heron was high in For protein, micro-Kjeldahl and a dye-binding yield and low in protein. technique are used. Lysine is determined mainly Feillet (1965) in his thesis on the influence of by autoanalyzer following acid hydrolysis; a genetic, agronomic and technological factors on modification of the Mossberg-Munck dye-binding the proteins of wheat described techniques for procedure for basic amino acids has also been fractionating wheat protein. By growing different used. Tryptophan is determined by a modified varieties under the same conditions, and the same ion exchange procedure. varieties under differing conditions, he concluded Large variations have been found among the that the albumin fraction is "specific" and may protein contents of wheats so far analyzed, levels be used to differentiate T. ru/gare from T. durum. ranging from 6 to 23% with a mean of 12.9%. The gliadin fraction is, he believes, "varietal" and Lysine per unit of protein ranged from 2.2 to characterizes each variety. The content of protein 4.2% with a mean of 3.2%. in flour is to a large extent influenced by environ­ Low protein wheats (mean of9.5% protein) and mental and agronomic factors, though the content high protein wheats (mean of 18.8% protein) dis­ of soluble protein appears to be heritable. play different amino acid profiles. Lysine per unit The influence of variety, soil and fertilizer of protein is negatively correlated with protein treatment on wheat yield, on protein and on other in low protein wheats but no similar correlation is nutrients was studied by El Gindy et al. (1957). found in high protein wheats. Lysine was positively Pawnee (hard red winter), Seneca (soft red winter) correlated with threonine, leucine and tyrosine, and Cornell 595 (white) wheat were grown on low­ and negatively correlated with valine; genetically fertility acid soil in 1951 and on well-limed fertile influenced increases in lysine should not therefore soil in 1956. Apart from soil characteristics, con­ be associated with a decrease in the threonine. ditions during the two years were much alike. When 39 wheats with the highest lysine values Protein in the wheat was affected by variety, were compared with 16 wheats with the highest soil and fertilizer treatment. The influence of each protein content to find which group provided the of these factors and the variety x soil interaction largest quantities of the essential amino acids per were highly significant but variety displayed much HULSE/LAING: PROTEINS OF TRITICALE 73

TABLE 45. Useful or potentially useful genetic sources of genes for high protein and/or high lysine in wheat (data from Johnson and Mattern 1972).

Selection Useful trait P.I. or C.I. no. Source High protein High lysine

Atlas 50 12534 USA x Atlas 66 12561 USA x Atlas x Comanche crosses USA x Frondoso 12078 Brazil x Frondoso derivative Purdue 28-2-1 USA x Aniversario 12578 Argentina x Aniversario-derived line NB66565 USA x Nebraska Male Fertility Restorer NB542437 USA x Nap Hal P.I. 176217 India x x Hume2 x Nb4 -Agrus-Tc7 SD69103 USA x April Bearded 7337 England x x Hybrid English 6225 England x x Pearl 3285 Sweden x (probable) Fultz x Hungarian 11849 USA x (probable) Fultz Sel.-Hungarian x Minturki-Fultz Sel. 12756 USA x (probable) Norin 10-Brevor, Sel. 14 x 27-15-Rio-Rex, Sel. 53 13447 USA x (possible) Norin 10-Brevor, Sel. 14 x Brevor Sib, 50-3 13449 USA x (possible) 22A 5484 USSR x (probable)

TABLE 46. Kjeldahl nitrogen content and arginine, lysine and valine content as percent of total nitrogen (data from Price 1950).

Nitrogen Arginine Lysine Valine Wheat {ofo) (Nitrogen% total nitrogen)

White English I. 58 10.2 4.12 3.56 New South Wales Australian I. 81 9.08 3.81 3.44 Plate 2.27 8.94 3.46 3.32 No. 2 Manitoban 2.43 8.75 3.39 3. 15

the largest variance and appeared to be the South Wales, Australian, (3) Plate and (4) a No. 2 dominant factor. Manitoban. He found that amino acid content tended to be higher in samples of lower Kjeldahl nitrogen content; this was particularly the case AMINO ACID COMPOSITION with arginine, lysine and valine (see Table 46). Price ( 1950) determined micro biologically the No similar trend was noted with phenylalanine, essential amino acid and tyrosine content of four histidine, tryptophan or tyrosine though none of samples of wheat: ( 1) White English, (2) New these amino acids showed a reverse tendency. 74 MONOGRAPH IDRC-02le

TABLE 47. Protein and lysine in six varieties of The protein content and percent lysine in the T. durum, (14% moisture basis) (data from Lawrence protein of the durum wheats Stewart, Mindum, et al. 1958). Langdon, Ramsey, Sentry and Pentad (red durum) are shown in Table 47. Lysine(% protein) varied Protein Lysine from 2.7% for Pentad to 3.3% for Stewart. N x 5.7 in protein The mean values for percent lysine in the protein Variety (%) (%) were: for 18 samples of other Triticum species, 3.0; for 13 samples of genera related to wheat, Stewart 13 .4 3.30 Mind um 12.6 3.04 3.3; for 15 hybrid selections of crosses of Agro­ Langdon 12.4 3.02 pyron elongatum with wheat, 3.1. Ramsey 13.0 2.93 McDermott and Pace (1960) determined the Sentry 13.8 2.85 amino acid composition (ion exchange chromatog­ Pentad (red durum) 12.6 2.70 raphy) of flours milled from a Manitoba wheat and from Hybrid 46 wheat of, respectively, Kjeldahl nitrogen 2.31 and 1.68%, dry matter Bendicenti et al. ( 1957) determined the nitrogen, basis. The amino acid composition of the two by micro-Kjeldahl, in the Italian soft wheat flours was generally similar except for lysine and varieties San Pastore (I. 73%), Funo (I. 95%), arginine which were both higher in the low protein Damiano (l.98%) and Mara (2.01%) and the hard Hybrid 46. The same inverse relationship between varieties, Cappello (1.99%) and Rossello (2.00%). content of nitrogen (N) and content of lysine was They also compared the soft varieties Tevere also found in the flours milled from Svenno (2.60%) and Mara (2.71 %), the hard variety (2.47% N), Bersee (1.85% N), Holdfast (1.44% N) Garigliano (2.62%) and two American varieties and Arletta (1.18% N). Cheyenne (2.78%) and Pawnee (2.84%). Sihlbom (1962) reported on the Kjeldahl nitro­ The nitrogen content of the first group ranged gen content and essential amino acid composition from 1.73 to 2.01% and of the second from 2.60 (Moore et al. 1958) of the winter wheats, Banco to 2.84%. The amino acid contents (determined and Odin and the spring wheat Svenno, grown in chromatographically; cystine by the method of different locations in Sweden. The results are given Schram et al. (1954) and tryptophan micro­ in Table 48. An inverse relationship between biologically) showed a consistently higher percen­ lysine content and total nitrogen is evidenced by tage oflysine and lower percentage of glutamic acid these results. Only one of the wheats examined, in the low nitrogen wheats compared to the higher Svenno from Hasslarp, had a protein content nitrogen wheats. There were no appreciable above the 13.5% level at which, according to differences in the amino acid composition of soft Lawrence et al. (1958), the inverse relationship and hard wheat varieties of the same nitrogen is no longer demonstrable. content. Simmonds (1962) reported on the variability in Lawrence et al. ( 1958) found that lysine content amino acid composition (determined by ion ex­ (determined microbiologically) varied from 2.46% change chromatography) of six Australian wheats in the protein (N x 5.7) to 3.84% in some 230 and the flours milled from them. The varieties, wheat varieties selected to be representative of origins, and Kjeldahl nitrogen as percent on dry commercial varieties. The overall mean lysine con­ weight were: Crossbred (Tichborne, NSW) 2.81 % ; tent on a percent protein basis was 2.89% for all Gabo (NSW) 3.14%; Charter (Allsopps NSW) wheats of about 13.5% or more protein; below this 2.84%; Charter (Boggabri, NSW) 2.74%; Brough­ level there was a highly significant inverse relation­ ton (Querindi, NSW) 1.67%; Sabre (Querindi, ship between protein content and lysine content as NSW) 1.57%. percent of protein. The wheat samples, although differing in nitro­ Three winter wheats, Rio, Brevor and Elmar, gen content and type, resembled one another in and three spring wheats, Marfed, ldaed, and Baart amino acid composition, and the results confirm grown at Pullman and Lind, Washington, and those of Price (1950), Gunthardt and McGinnis Moro, Oregon, in 1952, 1953 and 1954 showed no (1957), Lawrence et al. (1958) and McDermott significant difference in the lysine content of the and Pace (1960) all of whom observed an inverse protein. relation between total nitrogen and lysine content, HULSE/LAING: PROTEINS OF TRITICALE 75

TABLE 48. Nitrogen and essential amino acid content (excluding tryptophan) (in grams amino acid/ 16 g N) of Swedish winter wheats and the spring wheat Svenno (data from Sihlbom 1962).

Winter wheat Spring wheat Svenno

Banco. Banco. Banco Gist ad, Odin. Gist ad. Vaster-gard. Paarp. Ost ergot- Tierp. Tierp. Ostergot- Hasslarp. Hjortshoj. Skane Skane land Uppland Uppland land Skane Skane

Nitrogen °~ dry matter 1.99 1.46 I. 83 I. 84 2.41 2.32 2.91 2.34 Amino acids Threonine 2.66 3.29 2.89 2.84 2.77 2.76 2.66 2.59 Cysteine 2.20 2 .17 2.23 2. 16 2.25 2.09 2.03 I. 78 Valine 4. 32 4. 36 4. 36 4.65 4.13 4.40 4.24 4. 15 Methionine I. 51 I .62 1.61 I .63 I. 56 1.44 1.10 1.41 lsoleucine 3.49 3.55 3.05 3.38 3.65 3.62 3.64 3.89 Leucine 6.55 6.96 6.69 6. 17 6.62 6.47 6.31 6.81 Tyrosine 3. 13 3.32 2.92 3.08 2.97 3.03 3 .12 3.08 Phenylalanine 4.56 4.65 4.23 4.34 4.62 4.46 4.60 4. 56 Lysine 2.69 3.57 3. 14 2.88 2.77 2.50 2.60 2.56

expressed as percent nitrogen. in wheat. The lysine Shoup et al. ( 1966) determined (by auto­ contents found by Simmonds ( 1962) are higher analyzer) the amino acid composition (excluding than those found by McDermott and Pace, and tryptophan). of eight hard red winter wheat the amide. glutamic acid. phenylalanine. praline varieties and five hard red spring wheat varieties and serine contents are 10 to 15% lower. Simmonds composited from the 1963 crop grown in different (1962) suggests that this may be due to the areas of the United States. Single samples of different methods of hydrolysis employed. Seneca. a soft red winter wheat. Wells. a durum Hepburn and Bradley ( 1965) compared the wheat. and Omar. a soft white club wheat. were nitrogen (Kjeldahl-Gunning procedure) and the included. amino acid composition (microbiologically or by The histidine. arginine. threonine. glycine and ion exchange chromatography) of eight varieties methionine concentrations in the protein were of hard red winter wheat. four varieties of hard negatively correlated. and the glutamic acid and red spring wheat. one club wheat and one white praline contents positively correlated. with the wheat grown in different parts of the United total protein content of the wheats. As the protein States in the crop year 1960. The same variety in the wheat increased. the contents of the amino varied in nitrogen content according to location acids increased except for cystine and methionine. (see Table 49). Where the nitrogen content was the The proteins of the hard red spring wheats con­ same. the proportions of the amino acids were tained less lysine. arginine and methionine and nearly constant for all varieties. though cystine more cystine than did the hard red winter wheats. and methionine were somewhat higher in the spring Robinson and Sageman ( 1968) reported on the wheats. amino acid composition (by autoanalyzer) and Histidine. isoleucine and tyrosine varied least nitrogen content (by two described methods with nitrogen level. In the higher protein wheats which agreed exactly) of three South African glutamic acid. phenylalanine and praline tended wheats. Gondveld. Penkop and Wit Spitkop and to be higher and the remaining amino acids lower. three Australian wheats. Gabo. Gamenya and Hepburn and Bradley suggest the total amino acid Gala sown. on four sowing dates in April. May. contribution of any given sample of wheat is July and August under irrigation in the Ord determined primarily by the total amount of River Valley in North Western Australia. The protein it contains. total nitrogen (percent dry matter) varied from 76 MONOGRAPH IDRC-02le

TABLE 49. Nitrogen content of commercial wheats grown in various locations in the USA in the crop year 1960 (data from Hepburn and Bradley 1965).

Nitrogen (14% moisture basis) Type Variety Location (%)

Club El mar California I .41 White Baa rt Washington I. 93 Bison { Akron, Colorado 3.04 Alliance, Nebraska 2.37 N. Platte. Nebraska I. 98 Comanche { Akron, Colorado 2.98 Alliance. Nebraska 2.37 Garden City, Kansas I. 92 Concho { Akron, Colorado 2.97 Hard Red Winter Garden City, Kansas 2.37 N. Platte, Nebraska I. 76 Pawnee { Akron, Colorado 3.04 Alliance. Nebraska 2.40 N. Platte. Nebraska I. 97 Kaw Garden City, Kansas 2.37 Omaha Fort Collins, Colorado 2.42 Ottawa Fort Collins, Colorado 2.40 Tascosa Cherokee, Oklahoma 2.34 Thatcher { Williston, N. Dakota 3.34 Fargo, N. Dakota 2.35 Canthatch { Williston, N. Dakota 3 .13 Hard Red Spring Fargo, N. Dakota 2.26 Selkirk { Langdon, N. Dakota 2.42 Minot, N. Dakota 2.21 Conley Fargo, N. Dakota 2.43 Lee Minot. N. Dakota 2.44

3.86 (Gabo) to 3.38 (Gala) but no significant of grain were found to be positively and signifi­ difference in amino acid composition was cantly correlated with grain protein. A negative. demonstrated. but not statistically significant, relation was found Deosthale et al. ( 1969) found that the protein between lysine percent in the protein and protein content (Kjeldahl N x 5. 7) of 50 lines of wheat content. grown under identical conditions at the Indian Mehdi ( 1972) reported on the amino acid Agricultural Research Institute, New Delhi, varied composition of five wheat varieties, Sharbati from 8.6 to 16.9%. The lysine content (micro­ Sonora, CA-82, WL-2, Kalyan Sona and D49 I -5, biological assay) varied from 2.14 to 2.97 percent and I I strains, grown in 196 7-68 at the Indian of total protein. An inverse correlation was Agricultural Research Institute, New Delhi. The observed between total protein and lysine as method of determining the amino acids is not percent protein. stated nor is the method of determining protein. Gupta ( 1970) reported on the lysine and The highest lysine content was provided by methionine content (microbiological assay) in 17 Sharbati Sonora, 2.86 g/ I 00 g protein. Of the Indian and nine dwarf Mexican wheat varieties strains, HD(M)-1637, HD(M)-1653, HD(M)-1953, grown at the Indian Agricultural Research Insti­ HD(M)-1633, HD(M)-1638, HD(M)-1659, tute, New Delhi. Lysine and methionine as percent HD(M)-1592, HD-1622, HD-1728, HD(M)-1668 HULSE/LAING· PROTEINS OF TRITICALE 77 and HD-1632. the highest lysine contents were high unadjusted lysine values reflects the effect provided (g/100 g protein) by HD(M)-1668. of protein level upon lysine content (% P). 2.80. and HD(M)-1593. 2.73. HD(M)-1633 and Since these results. demonstrating wide varia­ HD-1728 were deficient in lysine. HD-1728 was tions in protein and lysine contents among the also deficient in methionine and cystine. Kalyan wheats of the World Collection. were all obtained Sona. HD(M)-1633 and DA 491-5 were highest in from grain produced at one location. Mesa. methionine which is the first limiting amino acid Arizona. USA. Johnson and Mattern consider it in diets of mixed cereals and pulses. reasonable to assume that a significant portion of Johnson and Mattern (1972) analyzed 12.681 the protein and lysine variation is genetic in origin. common wheats from the World Collection for However. only extensive trials over a period of protein (mostly by Kjeldahl) and lysine (mostly by years at several locations can firmly establish the ion exchange). Proteins are quoted on a dry weight genetic potential for high protein. Nevertheless basis (dwb) or at 14% moisture content (14 H20). the fact that lysine (% dm) is positively correlated and lysine as percent protein (% P) or as percent with protein content provides strong evidence that total dry matter (% dm). Protein (dwb) ranged the nutritional value of wheat can be increased from 6 to 23%. with a mean of 12.9%; lysine by increasing its protein content. (% P) ranged from 2.2 to 4.2% with a mean of Johnson and Mattern (1972) report that neither 3.2%. An overall negative relation between pro­ protein nor lysine content is influenced by kernel tein and lysine (% P) was established. In the 8 to size provided the kernels are plump and healthy. 14% protein range a strongly negative relation On the other hand. shrivelled kernels were higher between protein and lysine (% P) was evident. in protein and lysine than plump kernels.

This relation was less pronounced above 14° 0 protein and virtually disappeared above 16% BIOLOGICAL QUALITY protein. Bannerjee and Das ( 1964) compared 12 varieties Lysine (% dm) displayed a strongly positive of Indian wheat grown at the Indian Agricultural linear relation when plotted against protein. As Research Station. New Delhi. in feeding tests with protein increased from 8 to 21 % (dwb). lysine weanling albino rats. five in a group. over four (% dm) increases from 0.30 to 0.60%. weeks. fed ad libitum. The protein content (method Johnson and Mattern (1972) list: of determination and conversion factor not stated) (A) common wheats in the World Collection of the wheats varied from 13.45/;, (NP 809) to having highest protein (dwb) contents which 10° 0 (NP 797). Protein level in the diets varied range from 22.0 to 18.9%; lysine (% P) from 6.3° 0 for NP 797 to 8.61% (NP 823). The ranged from 2.42 to 3.14 and the lysine ratio weight gain: protein intake varied from 1.63 (% dm) from 0.44 to 0.66. to 1.96. There was no correlation among any of (B) common wheats having highest lysine (% P) the statistics quoted. contents which range from 4.26 to 3.83; Triumph 64 winter wheat. protein 15. 7~~ (Kjel­ protein contents (% dwb) range from 6.9 dahl N x 5.7. dry basis) and a commercial wheat. to 10.3. protein 10.8" 0 • were fed. on an equal weight of (C) durum wheats. from 3400 entries in the World grain basis. in trials with mice. five per group. Collection. having highest protein (% dwb) over 28 days; a 10% casein diet was included for which range from 21.3 to 19.4. lysine(% P) comparison. The higher protein Triumph 64 ranging from 2.57 to 3.09. and lysine(% dm) produced better weight gains and feed efficiency from 0.50 to 0.62. ratios (feed consumption: weight gain) than did (D) durum wheats with highest lysine (% P) the lower protein commercial wheat (Johnson and which range from 4.29 to 3.75. protein (% Mattern 1972). dwb) ranging from 7.3 to 12.1. They also list common and durum wheats with the highest adjusted lysine values. the negative Environmental Influences linear regression oflysine (% P) on protein(% dwb) being used to adjust lysine values to a common TOTAL PROTEIN protein level of 13.5% (dwb). The fact that fewer Pereira (1963) reported on the vanahons in wheats appeared with high adjusted than with protein content (method of determination not 78 MONOGRAPH IDRC-02le

TABLE 50. Grain yield and protein content (N x 5. 7 were marked effects with location in protein con­ dry matter basis) of seven varieties at high-yielding and tent of the grain. The highest levels were given by low-yielding nursery sites of the International Winter the Mexican dwarf wheats. e.g. Sonora 64, 14.2~ 0 • Wheat Performance Nursery grown in 1969 (taken Lerma Rojo, 13.8~/0 , Sonora 63, 13. 7%. from data presented by V. A. Johnson at 3rd FAO/ Upreti and Abrol ( 1971) reported on the pro­ Rockefeller Foundation Wheat Seminar, April 29- tein content (Kjeldahl N x 5.7) of 21 strains of May 13, 1970, Ankara, Turkey) (Johnson and Mattern 1972). triple gene dwarf wheats grown at Delhi, India during the crop season 1968-69. Protein contents

Kabul, Kermanshah, ranged from 10.4 to 15.5°10 • 18 of the wheats Afghanistan Iran having protein contents above 12.0%. The effect of location on the protein content of Yield Protein Yield Protein different varieties is illustrated by Table 50 which Variety (q/ha)" (%) (q/ha)• (%) shows the grain yield and protein content of seven varieties at the high-yielding site of the Inter­ Nursery mean 52 13.8 16 14.8 national Winter Wheat Performance Nursery at Yorkstar 66 10.4 13 13.5 Kabul, Afghanistan and at the low-yielding site at Heine VII 61 15.9 18 15.4 Kermanshah, Iran (Johnson and Mattern 1972). Bankuti 120 I 60 14. I 16 14.7 NB67730 53 17. I 14 15.9 Riley 67 50 14.5 13 14.7 Gaines 46 10.8 II 14.0 AMINO ACID COMPOSITION Atlas 66 44 16.5 13 18.5 McElroy et al. ( 1949) determined micro bio­ logically the content of nine essential amino acids •q/ha = quintal (I 00 kg)/ha. in Marquis wheat grown in nine different soil zones in Alberta, Canada. The soils were (a) gray­ stated) in the grain of 13 varieties of wheat grown wooded, (b) gray transitional, (c) black, (d) black­ in 10 localities in Portugal. Protein content varied transitional, (e) dark brown, (/) light brown. more within. the same variety grown in different Kjeldahl nitrogen of the wheat ranged from 1.94 locations than between different varieties grown in to 4.03%. with a mean of 3.02%. Lysine content the same locality. For example, Verdeal Rijo varied from 2.1 to 2.5% with a mean of 2.2% of varied from 15.9% (dry matter basis) at Braga to the protein (Kjeldahl N x 6.25). An inverse 9.8% at Elvas. a difference of 6.1 %. At Elvas. the relationship between total nitrogen and lysine as largest difference was 4.5% between Magueija, percent of total nitrogen was observed. 13.5% and Quaderna 9.0%. and at Braga between Swaminathan et al. ( 1969) reported on the effect Verdeal Rijo 15.9% and Quaderna 13.1%. Pereira on protein and lysine content of growing six also found that lysine in the protein varied in­ varieties of wheat at four locations in India. The versely with protein content. results are presented in Table 51. The lowest A survey of improved Indian and Mexican protein, 11.72% (Kjeldahl N x 5.7) was recorded dwarf wheats for protein content (Kjeldahl N x 5. 7) for S.308 (Sonalika) at Bijnoor. a low rainfall area was reported by Austin et al. (1970). Some 103 with a sandy soil. The highest value was 18.29% varieties were involved and 29 locations in the for C.273, a local tall wheat grown at Bhind on a five wheat zones of India in three crop years but clay soil. The lysine (microbiological assay) was not all the varieties were grown in all the locations. also influenced by location. At least nine observations were made on any one Eggum ( 1970) also compared the amino acid variety. content in grams per 16 g nitrogen, and nitrogen The varieties were divided into: content, and the protein value. expressed as true (i) low (less than I 0% protein). 7 varieties; digestibility, Biological Value, NPU, and utilizable (ii) medium (10 to 12% protein). 63 varieties; nitrogen (NPU x %N/100) of certain higher pro­ (iii) above 12% protein (ranging from 12.1 to tein wheats. The comparisons are given in Table 52. 14.2%). 33 varieties Sharbati-Sonora ranks highest as measured by The mean protein values for the locations varied Biological Value and NPU; Super X as measured from I 0.5% (range 9.2 to 11.6%) at Jaunpur to by true digestibility, and Jarral as measured by 16.4% (range 14.5 to 17.7%) at Bardoli. There utilizable nitrogen. HULSE/LAING: PROTEINS OF TRITICALE 79

TABLE 51. Effect of location on protein and lysine content in six varieties of wheat grown at four locations in India (data from Swaminathan et al. 1969).

Location

Delhi Bijnoor Bhind Sri-Ganga Nagar

Lysine Lysine Lysine Lysine Protein (% Protein (% Protein (% Protein (% Variety (%) protein) ('%,) protein) (%) protein) (%) protein)

NP.871 14.60 2.70 13 .12 3.09 18.00 2.25 13.40 2.73 C.273 15 .10 2.06 14.27 2.69 18.29 2.28 13.26 3.01 S.227 13.00 2.48 12.69 2.63 16.40 2.56 13.70 2.84 S.308 13.70 2.51 11. 72 3.32 16.84 2.58 12.68 2.76 Lerma Rojo 14.40 2.24 11. 92 2.80 16.93 3.41 12.60 2.86 PV.18 12.10 2.11 12. 13 2.43 16.40 2.50 11.96 3.58

TABLE 52. Amino acid content (grams/ 16 g N), nitrogen content and protein value(%) of high protein wheats (all wheats grown in Denmark except Sharbati-Sonora which was grown in India) (data from Eggum 1970).

Sharbati­ Species Sonora 64 Super X Inia Jarral Colibri Sonora Average Nationality Mexican Mexican Mexican Mexican German Indian Danish

Lysine 2.47 2.45 2.30 2.39 2.54 2.39 2.55 Methionine l. 66 l . 54 l . 49 l . 4 7 l . 49 l. 52 l. 82 Cystine 2.21 2.05 2.05 2.07 2.11 2.30 l. 81

Aspartic acid 4.48 4.37 4.51 4.43 4.92 4.95 5.26 Threonine 2 . 72 2 . 64 2 . 59 2 . 72 2 . 81 2.84 3.02 Serine 4.45 4.48 4.21 4.69 4.58 4.M 4.73 Glutamic acid 33.26 34.30 34.25 36.25 33.79 30.57 35. 77 Glycine 3 . 78 3 . 8 7 3 . 80 4. 15 3 . 99 3.98 4.19 Alanine 3. 17 3. 13 3. 04 3. 22 3 .40 3.36 3.77 Valine 4.01 4.07 4.05 4.17 4.31 4.13 4.58 Isoleucine 3.54 3.48 3.48 3.53 3.57 3.39 3.38 Leucine 6.47 6.53 6.28 6.67 6.67 6.33 6.79 Tyrosine 2. 44 3. 60 3 .47 3. 81 3 .49 3. l l 3 .14 Phenylalanine 4.74 4.83 4.87 4.88 4.74 4.40 4.41 Histidine 2. 27 2. 29 2. 30 2. 23 2. 38 2.09 2.29 Arginine 4.54 4.55 4.25 4.36 4.81 4.13 4.65 Tryptophan 1.37 l.ll 0.95 l.08 l.00 l.47 l. 12 N as % of dry matter 3.02 3.03 3.12 3.64 2.93 2.40 2.01

Protein value expressed as: True digestibility• 93.4 94.4 90.8 93.2 91.5 89. l 89.6 Biological valueb 52.0 52.0 49.4 49.4 55.0 60.5 59.0 Net protein utilization (NPU)b ~.4 ~.l M.3 ~.8 ~.3 53.9 52.9 Utilization nitrogen (NPU x % N/100) 1.46 l .49 1.38 1.67 l .47 l.29 l.06

•True digestibility is not defined in the paper but is generally regarded as the proportion of food nitrogen that is absorbed (National Academy of Sciences-National Research Council 1963). bThomas-Mitchell method. 80 MONOGRAPH IDRC-021e

TABLE 53. Average grain yield, protein content, and lysine content for varieties grown in the International Winter Wheat Performance Nursery in 1969 and 1970 (data from Johnson and Mattern 1972).

2-year 2-year 2-year average grain average protein average lysine yield content content

Variety (q/ha) (Rank) ("o) (Rank) ('/o) (Rank)

Winter varieties Bezostaia 42.4 13.8 25 2.9 8 Blue boy 36.4 4 14.1 21 3.0 3 San Pastore 33.9 14 14.1 21 2.9 8 Sturdy 36.4 6 14.8 II 2.8 21 Timwin 37.7 2 14.7 15 3.0 3 Parker 36.4 5 14.8 II 2.9 8 Fertodi 293 35.9 8 15. I 8 2.8 21 Benhur 34.2 12 15. I 8 2.8 21 Scout 66 36.6 3 14.5 18 2.9 8 Yung Kwang 34.2 13 14.8 II 2.9 8 Arthur 36. I 7 14.7 15 2.9 8 Gage 34.6 10 14.8 II 2.9 8 Stadler 34.4 II 13.7 26 3.0 3 Heine VII 33.9 15 14.7 15 2.9 8 Lancer 34.7 9 14.2 20 2.9 8 Shawnee 33.4 17 14.1 21 2.9 8 Riley 67 33.9 16 14.5 18 3.0 3 Yorkstar 33.4 18 12.8 28 3. I Bankuti 120 I 32.0 20 15.5 5 2.8 21 Triumph 64 32.8 19 15.2 7 2.9 8 NB67730 31. 5 21 16.9 3 2.8 21 Atlas 66 30.8 22 17.9 2.7 28 Purdue 28-2-1 30.0 24 17.5 2 2.8 21 Winalta 30.2 23 14. I 21 2.9 8 Cappelle Desprez 29.0 25 16.0 4 2.8 21 Gaines 28.7 26 13.4 27 3. I Felix 27.0 27 15. I 8 2.9 8 Odin 24.6 28 15.5 5 3.0 3 Spring varieties Lerma Rojo 64 32.4 14.5 (1969 only) 2.8 (1969 only) INIA66 25.5 14. 5 (1969 only) 2. 8 (1969 only)

Twenty-eight winter wheat vaneties and two were used for determining the protein. The Russian spring wheat varieties were grown at the Inter­ wheat Bezostaia with a UDY protein mean of national Winter. Wheat Performance Nursery, 13.2% and a lysine mean of 2.8% compared well which has sites throughout the world, in 1969 and with the other varieties and ranked first in yield 1970. The 2-year average grain yield, protein (see Table 53). (Johnson and Mattern 1972). content, and lysine content of these varieties, with A selection of wheat varieties with high lysine their respective rankings, are summarized in values when grown at Mesa, Arizona, were re­ Table 53 (Johnson and Mattern 1972). grown at other sites in the United States. The The protein, and lysine content (by auto­ results indicated that in terms of lysine most of the analyzer), mean and range, for NB 67730, Triumph varieties were not genetically different. However, 64, Bezostaia and Sturdy grown at 15 locations the spring varieties Pl 7337 and Pl 5484 showed are given in Table 54. A modification of the UDY promise, and the winter varieties Atlas 66, Pl dye-binding procedure and/or the Kjeldahl method 135067, Pl 135070, Pl 166913, Pl 94526, Pl HULSE/LAING: PROTEINS OF TRITICALE 81

TABLE 54. Protein and lysine contents (range in parentheses) for four wheat varieties from 15 locations in the First International Winter Wheat Performance Nursery (data from Johnson and Mattern 1972).

Lysine

In UDY In Kjeldahl UDY protein Kjeldahl protein protein protein 0 0 ( (o~) ( 0 sample) 0 sample) (/o)

NB 67730 16.0 2.7 (11.4-21.9) (2. 3-3. I) Triumph 64 14. 7 2.7 (I I. 3-18 .6) (2. 5-3. I) Bezostaia 13.2 2.8 ( 9.8-15.2) (2.6-3.2) Sturdy 14.0 14.6 2.8 2.7 (not available) (10. 7-18.3) (not available) (2 .4-3. 9)

173438. Pl 11680. Cl 13449 and Pl 117018 had Vauxhall and Burdett in Southern Alberta, higher protein and lysine contents than Triumph Canada. 64 (Johnson and Mattern 1972). Hutcheon and Paul ( 1966) reported on growth chamber experiments to control the protein con­ tent of Thatcher wheat by nitrogen fertilization Agronomic Influences and moisture stress and found that it was possible TOT AL PROTEIN to increase both yield and protein content at the

In general. chemical fertilizers have not been same time in the protein range of 11 to 16°·0 . widely used in the past in the major wheat grow­ When protein contents above I 6'j0 were obtained, ing and wheat exporting areas of the world but it was at the expense of yield. only where intensive cultivation is practised. and The nitrogen nutrition and yield relations of with the object of increasing yield (Moran 1959). Nugaines. a semi-dwarf high-yielding soft white The influence of chemical fertilization has winter wheat, was studied by Laopirojana et al. received some attention in India where the effect ( 1972) at Oregon Agricultural Experimental of moisture regimes and fertilization practices on Station. Corvallis. High nitrogen fertilization wheat yield and protein in the grain has been increased grain protein percentage. studied by Singh and Prasad (1966). Reddy et al. Grain and plant nitrogen relations were studied ( 1968). Panwar et al. ( 1971 ). Sharma et al. ( 1972). in eight spring wheat crosses seeded at three Singh and Dastane ( 1971 ). locations in Montana. USA. Grain nitrogen In the United Arab Republic, Barakat et al. content was found to be negatively correlated to ( 1970) have reported on the interaction between grain yield and to the grain: straw ratio (Mc Neal soil salinity and nitrogen fertilizer on the yield and et al. 1972). protein content ofa 145 Giza wheat variety. Six Indian wheats and 14 wheats from overseas Beech and Norman ( 1966). Williams ( 1966) and were grown at Durgapura, Jaipur. India under Taylor and Gilmour ( 1971) have reported on the similar conditions and with three levels of nitrogen effects of climatic factors and agronomic practices fertilization (67.2 kg. 134.5 kg and 201.7 kg/ha). on the yield and protein content of wheats grown The wheats were EG 953. EG 954 and EG 144-0 in Australia. from Egypt; Charter and Gaba from Australia; Dubetz ( 1972) has reported on the effects of 184.P.2.A.l.E. and E.220 from Kenya; Cometo various levels of nitrogen fertilizers on the yield Semiduro from Brazil; Trigo Centeira from South and protein content of a bread wheat, Manitou. America; S.2414, S.54723. S.55362 and S.54887 and a semi-dwarf high yielding wheat. Pitic 62, from Mexico; C.286 and C.591 from the Punjab; grown under irrigation in 1969 and 1970 at Hy 65 from Madhya Pradesh; NP 790 and NP 825 82 MONOGRAPH IDRC-021e from the Indian Agricultural Research Institute; Pl 5, Kl 5 during pre-sowing cultivation and N20 and R.S. 31-l from Rajasthan. during early spring feeding. Under these con­ Protein (Kjeldahl N x 5.7), was determined and ditions total nitrogen, dry matter basis, was 3. l 2°~ crude gluten content (method of Austin and Miri following peas and 2. 70% following lupins in l 964, l 96 l) and kernel hardness (by a grain-hardness and 2.25% following peas and 2. l 3°~, following testing apparatus) were assessed. In all the varieties lupins in 1965. All nitrogen levels were higher in the percentage of crude gluten varied directly l 964 than in l 965. Fertilization increased the with the percentage of crude protein. Kernel hard­ nitrogen content of the albumins, globulins, ness was inversely related to gluten and seed weight. gliadins and gluten ins in the grain over the control. Varietal differences were highly significant. In Using pot trials in climatically controlled general, higher doses of nitrogen increased the growth chambers, Partridge and Shaykewich protein and gluten content in the grains and ( l 972) studied the effects of nitrogen, temperature decreased the hardness of the kernel but the and moisture regime on the yield and protein varieties did not all respond in the same way to (Kjeldahl N x 5.7) of the hard red spring bread fertilizer. Some varieties showed a decrease in wheat Neepawa. Photoperiod was l 6 hours light protein content when fertilizer level was raised and 8 hours dark at all times. above l 34.5 kg, e.g. Trigo Centeira, others At rates up to 100 ppm of nitrogen, grain yield showed an increase only at the· highest fertilizer increased with succeeding increments of fertilizer level. The mean protein ranged from l 2.0% for nitrogen. There was a variable response to the the lowest to l 2.4% for the medium to 13. !% final increment of 200 ppm, probably as a result for the highest level of fertilization (Gandhi and of phosphorus deficiency. Nathawat 1968). Percent protein was strongly influenced by nitro­ As the level of nitrogen fertilizer applied to gen and temperature treatments through their six unnamed varieties of wheat increased (0, 40, effect on yield. There was a significant negative 80, l 20, l 60 and 200 kg/ha) the protein content correlation between percent protein and grain (Kjeldahl N x 5. 7) also increased from 13. 7% (no yield. Significant increases in percent protein over fertilizer) to 16.9% (200 kg/ha) (Swaminathan et zero nitrogen treatment, resulted from 200 ppm al. 1969). nitrogen. Increased temperatures significantly de­ Foliar sprays of urea and parazate, individually creased grain yield in most instances and, indirectly, and in combination. increased the protein content increased percent protein. Moisture regime had no in NP 4, NP 52, NP l 65, NP 718, and NP 825 significant influence on grain yield nor a significant wheat varieties, but the varieties differed in their direct effect on percent protein. response (Swaminathan et al. l 969). A high protein wheat, Cl 14016 (protein

Ermolaev (l 970) studied the effects of the rate Kjeldahl N x 5.7 dry weight basis, 12.5° 0 ) and an of sowing, the time of sowing and the level of ordinary variety, Lancer (protein 10.8°,~) were fertilization on the properties of Bezostaia l grown over three years at IO sites in Nebraska wheat grown over four years near Russe in and with different levels of soil fertility. Protein Bulgaria. The amount of crude protein (N x 6.25) content increased in linear relation to nitrogen and dry gluten in the flour was most affected by fertilizer application. The high protein wheat the level of fertilization. The biggest effects were maintained its advantage over Lancer throughout obtained with 140 kg/ha N active substance, all rates of nitrogen application from 0 to l 20

100 kg/ha P20 5 and 60 kg/ha K2 0, at which level lb/acre (0 to l 34 kg/ha) demonstrating that the the grain yield doubled. Total crude protein in­ difference in protein content between varieties is creased by 19% and dry gluten by 44.3% com­ genetically based. Wheat of the high grain protein pared to the unfertilized control. ( 14 to l 7%) can therefore be obtained from high Mironovskaya 264 winter wheat grown in the protein strains grown under conditions of high Ukraine in l 964 and l 965, as part of a crop fertility (Johnson and Mattern l 972). rotation system, contained higher total nitrogen levels when following peas than when following lupins grown for forage (Vlasyuk and Kurinnaya AMINO ACID COMPOSITION l 97 l ). The best results were obtained using Gunthardt and McGinnis (I 957) compared the applications of P60, K90 at the ploughing stage, amino acid composition of four low protein HULSE/LAING: PROTEINS OF TRITICALE 83

) TABLE (I 0.3~ 0 ) and four high protein ( 16.5° 0 samples of 55. Protein content and lysine as percent Idaed wheat. The high protein samples were protein in five wheat varieties grown in Hungary in produced by a combination of nitrogen ( 120 lb. 1961 (data from Lindner 1964). ammonium nitrate/acre: 134 kg/ha) and water applications at the boot. flowering or milk stages Protein of growth: the low protein samples received from (N x 5.8) Lysine 0 to 40 lb nitrogen (0 to 45 kg) at seeding time (%) (%protein) only. The high protein samples contained signifi­ Hungarian cantly less lysine (microbiological assay) expressed Bankuti 1201 11.54 3.2 as lysine percent crude protein than did the low Russian protein samples. The lower lysine arose when Bezostaia 12.99 3.2 nitrogen applications were as late as the boot. Skorospelka 13.74 2.7 flowering or milk stages of growth. The lysine Italian content appeared to vary with the year. or yield. San Pastore 12.24 2.7 or both. Other changes in amino acid composition Autonomia 12.47 2.7 were not statistically significant. Large differences in protein content were produced by nitrogen fertilization but only slight differences in pro­ in 1961. The protein (N x 5.8) and lysine as per­ portional amino acid content of total crude cent protein are shown in Table 55. protein. Of the varieties studied. Bezostaia had the Sosulski et al. ( 1963) found that reduced water highest nutritional value as measured by the EAA supply. nitrogen fertilization. and higher air index of Oser ( 1951); Bezostaia and Bankuti 1201 temperatures increased the protein content of rated equally by the chemical score method of Thatcher wheat grown under controlled environ­ Mitchell and Block (1946). mental conditions in growth chambers. At higher Larsen and Nielsen ( 1966) determined nitrogen temperatures the soil moisture conditions exerted (micro-Kjeldahl) and amino acid content (auto­ the greater effect on protein content while the analyzer) of Koga wheat grown in pots ina mixture largest responses to nitrogen fertilization were of one part clay soil and two parts sand. with a obtained at the medium moisture level. The amino basal dressing and nitrogen as ammonium sulphate acid distribution was determined by the method of added at levels ranging from 0.025 g N/pot to Moore and Stein (1954a. b); cystine and trypto­ 5.0 g N/pot. phan were not determined. Their results were in agreement with those of Protein content ranged from I 0.2 to 23.0'\ other investigators. Increasing applications of (American Association of Cereal Chemists 1957) nitrogen to wheat caused an increase. in grams expressed on a 14° 0 moisture basis. for the samples amino acid nitrogen per I 00 g total nitrogen. in grown at three moisture and five fertility levels. glutamic acid. from 15.6 to 17.4 and praline. from Regression and correlation coefficients were cal­ 6.4 to 7.8 in the grain and a decrease in the lysine. culated between grain protein content and con­ from 3.6 to 2.6. and arginine. from 11.25 to 9.0. centration of each amino acid. Glutamic acid and They report small increases in ammonia and praline were positively correlated with increasing phenylalanine. and small decreases in aspartic acid. protein content: alanine. arginine. aspartic acid. threonine. glycine. alanine and valine. lysine. serine. threonine. and valine were negatively Ewald and Wenzel ( 1967) examined the effect correlated with protein content. Glutamic acid on protein composition and amino acid content and arginine were the principal amino acids (by column chromatography) on Opal summer affected by variations in protein content. wheat. sown on loamy soil. of nitrogen fertilization. Lindner ( 1964) examined the amino acid com­ As nitrogen fertilization increased. yield increased. position (Lindner 1956) of two Russian wheats, first at a higher rate then at a lower rate: grain Bezostaia and Skorospelka. two Italian. San nitrogen increased. first slightly then. at the higher

• Pastore and Autonomia. and one Hungarian levels of nitrogen fertilization. from 1.68 to l .86° 0 variety. Bankuti 120 I. all grown at the Agricultural i.e. about 11 %- Amino acid content as percent Research Institute at Martonvasar. Hungary. all total protein changed little with nitrogen fertiliza­ given similar fertilization treatment and harvested tion. the major increase being in glutamic acid; 84 MONOGRAPH IDRC-021e lysine showed no change. The results of pig feeding followed. The 2-year rotation. without fertilizer. trials conducted at another institute. and more produced a total essential amino acid content of fully reported by Brune and Thier ( l 968a.b ). 34.3 (lysine 3.4) and the single crop. without showed that at a fixed nitrogen intake the biological fertilizer. 32.6 (lysine 3.0). value for pigs was reduced as the nitrogen content Srivastava et al. ( 1971) studied the effect of increased with fertilization. nitrogen applied as urea at levels of 0. 20. 40. 60 Abrol et al. (1971) have reported on the effect and 80 kg N/ha on the protein (Kjeldahl N x 5.7) of soil fertilizer levels on the protein (Association and lysine and tryptophan in the protein. of S-227 of Official Agricultural Chemists 1960) and amino wheat grown under rain-fed conditions at the acid composition (autoanalyzer: tryptophan Indian Agricultural Research Institute. New Delhi. colorimetrically) of two dwarf wheats HD(M) In soil treatments the whole dose was applied 1592 and HD(M) 1620A grown in India. The before seeding: in foliar treatments. half was fertilizer levels were (a) high. N 134. P68 and K45 applied to the soil before seeding and the rest kg/ha and (b) low. N34. P20 and KO kg/ha. sprayed as a 3° 0 urea solution. In each spray IO Protein was fractionated by the method of Nagy kg N/ha was applied. the first spray at the post­ et al. (1941). tillering stage. and then. according to the treat­ At high fertilizer levels the percentage of protein ment. spraying was repeated at 7-day intervals. increased but the increase in HD(M) 1620A was Table 56 shows the effect of soil and foliar applica­ 38°~ while the increase in HD(M) 1592 was 14° ()" tions of urea on yield. protein. lysine as percent of In HD(M) 1620A the prolamin and glutelin protein (enzymatic decarboxylase method. Naik fractions increased by 51 ~~ and 26° 0 respectively. 1968) and tryptophan as percent of protein in HD(M) 1592. the prolamin and glutelin fractions (colorimetric method of Spies and Chambers increased by 34° 0 and 21 % respectively. The higher 1949). The highest lysine content. 3.22° 0 of the fertilizer level had little effect upon the albumin protein. was obtained as 40 kgN/ha applied to the and globulin fractions: consistent with other soil; the highest tryptophan. 1.45".o of protein. reported results. the increase in total protein was was obtained with soil plus foliar treatment at predominantly in the prolamin and glutelin 80 kg N/ha. fractions. The amino acid composition (by autoanalyzer) In HD(M) 1592 the high fertilizer level increased of a sample of S-227 wheat showing the highest the glutamic acid. proline. phenylalanine and protein percentage. I 0.43°~. obtained from soil leucine contents and decreased the lysine. valine plus foliar treatment at 80 kg N/ha was compared and threonine contents calculated as percent with one showing the lowest. 7.23° 0 protein from protein. In HD(M) 1620A the high fertilizer level 0 kg N/ha: lysine percent in the protein was in­ increased the glutamic acid. leucine and proline. versely correlated with total protein in the wheat. and decreased the threonine. cystine. isoleucine Mamchenkov and Platonov ( 1971) found that and tyrosine; lysine was scarcely reduced. as the protein content (N x 5. 7) of Mironovskaya The effect of fertilization and crop rotation on 808 winter wheat increased through the use of the Kjeldahl nitrogen and essential amino acid nitrogen. phosphorus and potassium fertilizers composition (determined chromatographically) of combined with crop rotation. the percent amino wheat and maize grown on reddish-brown forest acid composition in the protein (determined by soil from Saftica in Roumania was reported by paper chromatography) altered as follows: lysine. Dinca et al. ( 1971 ). In a single wheat cropping histidine. threonine and glycine decreased; cystine. system. high chemical fertilizer rates reduced the aspartic acid. glutamic acid. serine. tyrosine. content of the essential amino acids. Without phenylalanine. and leucine increased; alanine. fertilizers. better results were obtained with short valine and methionine did not change. rotations of two to four years. The best results in Elonen et al. ( 1972) studied the effects on spring terms of essential amino acid content were wheats of irrigation and nitrogen fertilization. obtained with fertilizer N48. P32 and a 2-year separately and together. in five field trials in rotation. The essential amino acid content (grams Finland in the years 1967 through 1970. The amino acid/16 g N) was then 54.6 (lysine 3.3) varieties were Svenno in trials I and 2. Norrona compared to 31. I (lysine 2. 7) when the same in trial 3 and Ruso in trials 4 and 5. The soil was fertilizer was used and single crop cultivation was a silty clay with an average pH of 5.6. containing HULSE/LAING: PROTEINS OF TRITICALE 85

TABLE 56. Effect of soil and foliar application of urea on yield. protein (N x 5.7). lysine and tryptophan content of ·s 227' wheat (data from Srivastava et al. 1971 ).

Yield of Protein Lysine Tryptophan Nitrogen Method of grain (g/JOOg (g/JOOg (g/JOOg (kg;ha) application (kg; ha) grain) protein) protein)

0 Control 1.812 8.00 2.50 I .03 20 Soil 2.373 8.09 2.73 1.10 40 Soil 3, 146 8.04 3.22 I .17 60 Soil 3,681 8. 72 2.96 1.14 80 Soil 3.736 9.10 3. 10 1.24 20 Soil + foliar 3,022 8. 14 2.23 I. 12 40 Soil+ foliar 3.295 8.86 2.73 1.09 60 Soil + foliar 3.582 9.35 2.29 1.24 80 Soil+ foliar 3.765 9.64 2.38 I .45 SE rates ±0.122 ±0.154 ±0.0565 SE methods ±0.061 ±0.109 ±0.04

about 4° 0 of organic carbon in the top soil; the Khera et al. ( 1972) examined the relation subsoils were heavy clay. The basal fertilizer between the phosphorus level in the soil and the dressing on the N 1 plots contained on average. protein quality of the Kalyan Sona wheat grown 68 kg N. 53 kg P and 81 kg K per hectare and was on it as measured by the lysine, methionine and placed in rows at a depth of 8 cm. Additional tryptophan contents of the grain. Kalyan Sona was nitrogen on the N 2 plots averaged 76 kg/ha and grown in pots each containing non-saline soil. of was applied as a surface dressing for the shoots as pH 7.0 to 8.4. sandy loam to clay loam in texture calcium nitrate in trials 1 and 2. as urea in trial 3. and low in organic carbon. To each 4.5 kg soil a or at a depth of 8 cm in connection with sowing basal dose of 89 ppm N. 37 ppm Kand adequate either as urea or ammonium nitrate limestone in amounts of trace elements were added. The method trials 4 and 5. of Cate and Nelson ( 1965) was used to correlate The grains were milled in a Quadrumat Junior the soil test analyses with the plant response data. to a 60 to 65";, extraction flour. Irrigation increased Under these conditions it was found that pro­ average yields by 1200 kg/ha. Protein (Kjeldahl tein quality. as measured by protein (Kjeldahl N x 5. 7) decreased with irrigation but this decrease N x 5.7) percent in the grain, and lysine (by could be offset by additional nitrogen fertilization. microbiological assay). methionine and trypto­ Irrigation significantly increased the proportions phan (by colorimetric methods) in the nitrogen of of lysine and isoleucine (by autoanalyzer) as per­ the grain was inferior when the available phos- cent of protein but the decrease in total protein content gave rise to a net decrease in lysine content. TABLE 57. Effect of additional nitrogen fertilization G!utamic acid increased while lysine and arginine (N 1 and N2 ) and irrigation on grain yield. protein declined with nitrogen fertilization but as total (N x 5.7) lysine in flour and in protein of flour. from protein was increased a net increase. on average Finnish wheats (data from Elonen et al. 1972). 8.3 ± 5.1°~. was manifested. Table 57 summarizes the position. Without With By increasing fertilizer nitrogen from 68 to 144 irrigation irrigation kg/ha and by irrigating twice in June. grain yields NI Nz NI Nz increased by 65° 0 without any noticeable changes in flour protein or lysine contents. Grain yield. kg/ha 2850 3210 3820 4690 There were no significant differences between Protein in grain. % 14. 31 16.56 12.20 14.43 the amino acid composition of wheat fertilized by Protein in flour.% 13.50 15.93 11 .25 13.54 urea or by ammonium nitrate limestone but Lysine in flour. because of the relatively small quantity of material mg/100 g 292 315 260 283 available. the confidence limits were wide. 86 MONOGRAPH IDRC-021e phorus levels were below 12.5 ppm. Below this Chambers 1949) as percent protein was signifi­ level phosphatic fertilizer is required if grain cantly decreased by inclusion of stover and weight and protein quality are not to suffer. following cowpeas but was unchanged following At Pahlavi University, Shiraz, Iran, the effect berseem. Rotation produced no significant of potassium and nitrogen applied to the soil, on differences in the albumin, globulin, prolamin and the lysine, methionine and total protein of Roush an glutenin fractions (modified Mendel-Osborn bread wheat was studied by Hojjati and Maleki method), albumins and globulins accounting for (1972). The soil was calcareous silty clay loam, about 40% of total protein. A complete amino initially high in potassium which was added at the acid analysis (autoanalyzer) as grams per I 00 g rates of 0, 25, 50 and I 00 kg K/ha in the form of protein, showed that aspartic acid, glutamic acid potassium sulphate under four levels of nitrogen, and praline were slightly higher, and lysine, 0, 50, I 00 and 200 kg N/ha as urea. All the potas­ histidine and arginine slightly lower, in the sium and half the nitrogen was applied at seeding highest protein sample (16.27%) than in the lowest in November 1967 and the rest of the nitrogen in protein sample (14.18%). April 1968. The field was irrigated before seeding Chlorophenoxy herbicides, particularly 2,4- and during the growing season. dichloro-phenoxyacetic acid (2.4-D) arc used to Yield of grain and total dry matter production control broad-leaved weeds in wheat and one of were not affected by potassium applications. Yield the effects which may occur is depletion of the was not affected by nitrogen up to 100 kg N/ha starch and sugars and a higher percentage of but decreased at the 200 kg level. The ratio of protein in the grain (Klingman 1953 and Shaw et al. grain to total dry matter decreased consistently 1955 cited by Pellett and Saghir 1971 ). Samples of with increasing application of nitrogen showing Najah wheat grown in the Lebanon and treated that it stimulated vegetative growth more than it at 2, 4 and 8 kg/ha at the jointing stage of growth did seed production. with the butoxyethyl ester of 2,4-D were com­ Nitrogen fertilizer consistently increased the pared with others from unweeded and hand­ protein content (micro-Kjeldahl N x 5.83) of the weeded plots. No significant differences were grain, from about 10% to about 14%; an approxi­ found in amino acid composition (by ion exchange mate increase of I% was recorded with each chromatography) between the grain from the un­ additional 50 kg N/ha. Lysine and methionine (by weeded and the hand-weeded plots. A slight microbiological assay) increased as percent grain; decrease in threonine and arginine and an in­ lysine as percent protein was negatively correlated crease in the praline content of the protein were with the protein content and with the N applied, observed in grain from the treated plot. High but positively correlated with K. Methionine as doses of 2,4-D may increase protein percentage percent protein did not change with varying rates but have only a minor effect on amino acid com­ ofN or K. position of wheat (Pellett and Saghir 1971 ). Pahwa et al. ( 1972) found that the inclusion of cowpeas (Vigna sinensis Savi ex Hassk) and berseem (Trifolium alexandrinum Juslen) in the BIOLOGICAL QUALITY crop rotation significantly improved the yield from Greer and Grindley ( 1954) reported feeding about 31 to 35 q/ha, of Sonora 64 bread wheat trials in which weanling rats of both sexes were fed harvested at New Delhi in 1968. The addition of ad libitum on diets in which whole ground winter maize (Zea mays) stover in the soil before the wheat grown in the United Kingdom provided all wheat crops decreased yield but increased calcium of the protein. The mean grain nitrogen contents in the wheat flour. Crude protein content in the were: (I) 1.81 % Ni dry basis for grain which had grain (Association of Official Agricultural received no spring dressing, (2) 1.92% Ni dry Chemists 1965) increased following cowpeas and basis for grain which had received late April berseem and decreased when maize stover was dressing (2 cwt nitro chalk per acre), (3) 2.18% added but these differences were not significant. Ni dry basis for grain which had received the late Lysine (Naik 1968), methionine and cysteine April dressing plus an equivalent dressing in late (autoanalyzer) as grams per 100 g protein were May. Three feeding trials were conducted, with not significantly different; tryptophan (Spies and variations in the oil and vitamin supplementation HULSE/LAING: PROTEINS OF TRITICALE 87 to the diets. In two trials 10 pairs of rats were used mer wheat Opal grown in 1964 at the same and in the third, eight pairs. The mean values for institute. The Opal wheat is that referred to by nitrogen intake, dry basis, were: (a) no spring Ewald and Wenzel ( 1967). The fertilization con­ dressing 0.121 g N2 per day; (b) spring dressing ditions for Heine VI I were 40 kg/ha P2 0 5 and 80

0.150 g N2 per day. In all three experiments the kg/ha K 2 0 and nitrogen varied to give 0, 30, 60 growth of the rats receiving the spring dressed and 90 kg/ha. The conditions under which the wheat exceeded that of the rats consuming the two wheats were grown are not therefore com­ wheat which had received no spring dressing. parable. Growth increment, grams per grams nitrogen in­ The method of amino acid analysis is not stated take, was: no spring dressing 9.09; spring dressing but since the Heine VII and the Opal were grown 8.67 or a PER (protein N x 5.7) of: no spring at the same institute, although in different years, dressing 1.58, spring dressing 1.52. Greer and the amino acid analysis method of the Heine VII Grindley conclude that the improvement in grain may have been the same as that used for Opal by nitrogen brought about by spring dressing repre­ Ewald and Wenzel (1967). A comparison of the sented a protein increase of normal nutritive value. amino acid content as percent protein showed no Bains ( 1953) studied the effect of nitrogen (60 or very slight differences in lysine in response to lb N 2/acre; 67 kg/ha) and phosphorus (25 lb fertilization. Leucine, isoleucine and phenylalanine

P2 0 5/acre; 28 kg/ha) on the nutritive value of were lower in the winter wheat Heine VII with C409 wheat at Punjab Agricultural College and nitrogen fertilization but were higher in summer Research Institute, Lyallpur. The fertilizer treat­ wheat. Arginine was higher in winter wheat and ment consisted of (I) green manure ( Caianus lower in summer wheat with fertilization. Threo­ indicus) 5 tons/acre ( 12,544 kg/ha), (2) potassium nine increased in summer wheat with fertilization nitrate, (3) potassium nitrate with green manure, but remained the same in winter wheat. There ( 4) superphosphate, ( 5) superphosphate with green were no significant changes in amino acid com­ manure, (6) superphosphate with ammonium position of either wheat with fertilization. sulphate, (7) superphosphate with ammonium No correlation was evident between the amino sulphate and green manure. All the fertilizer acid scores of the grain (Mitchell and Block 1946; treatments improved yield per acre. Treatments Oser 1951) and the results of feeding trials with (I), (2), (3), (6) and (7) increased, and treatments pigs and rats. The winter wheat and the summer ( 4) and (5) decreased protein (N x 5. 7) in the grain. wheat could not be compared with one another However, all treatments improved output of because of the different conditions under which protein per acre, the most productive being treat­ they had been grown but both wheats grown under ment (7) with almost double the unfertilized con­ high levels of nitrogen fertilizer produced statisti­ trol, followed, in descending order, by (5), (2), cally significantly reduced "productive protein (6), ( 4), (3), and (I). values" (apparent NPU: nitrogen balance as The biological value of the grain was examined percentage of nitrogen intake) in pigs compared in rat feeding trials using modifications of the to the wheat grown without nitrogen fertilization. "balance sheet" method of Mitchell ( 1925). The A modified Thomas-Mitchell method for nitrogen diets, fed to provide 5% protein from the wheat metabolism in pigs was used. The difference was were fed to adult male albino rats, six per group, smaller but still significant with young rats for a 7-day period following seven days on a (initial weight 40 g, 10 per group) but was not seen nitrogen-free diet. The nitrogen-free diet was also in rats weighing 60 g (Brune et al. 1968). fed at the end of the trial. Under these conditions, Ferrel et al. ( 1970) report experiments in which that is when the test samples were fed at the same wheat, fortified with widely different levels of protein level of 5%. there was no noticeable effect lysine, was stored under several conditions. of the variations in the protein content of the Subsequently PERs were determined at a con­ samples resulting from fertilization. stant lysine level. It is difficult to draw clear con­ Brune and Thier (I 968a, b) examined samples clusions from the reported results through Ferrel of the winter wheat Heine VII grown in 1961 at et al. suggest that the lysine naturally present in the lnstitut fur Pflanzenernahrungslehre und the wheat suffered greater change than the added Bodenbiologie, Stuttgart/Hohenheim and the sum- lysine. 88 MONOGRAPH IORC-02le

Wheat and Rye Compared stated) of whole hard wheat flour, protein 14. 7%,

whole rye (pumpernickel) flour, protein 9.44/0 , Genetic and Varietal Effects and Arkansas (USA) grown whole rye flour of 10.1 % protein in feeding experiments with 12 Jones et al. (1948) reviewing the comparative male and 12 female albino rats per group, over 1O growth-promoting (rate of weight gain) values for weeks. The cereals provided the only source of rats, of the proteins of cereal grains, commented protein in the diets, the protein being fed at 9%, that compared with the proteins of most cereal 8% and 5% levels. At all levels of protein intake, grains, rye had received comparatively little rye protein was superior to wheat protein in attention. In their own experimental work they terms of weight gain and ratio of weight gain: compared whole rye, protein content 10.96/ 0 protein intake. (N x 5.83) with other cereals including hard spring A paired feeding experiment was also conducted wheat, protein 14.24% (N x 5.83) and soft winter over 35 days with the protein at the 8% level. On wheat, protein 11.02% (N x 5.83) as the sole the same amount of food and of protein daily, the source of the protein in rat diets. In two sets of rye-fed animals gained 70.4% more in body weight experiments wheat and rye diets were compared than those on the wheat flour. at levels of 4.5%, 7.5% and 9.5% protein. The de Vuyst et al. ( 1958) determined the amino results are shown in Table 58. acid composition of various grains grown in The superiority of rye protein over wheat pro­ different locations in Belgium and intended for tein found in these experiments confirmed the animal feed. The protein contents (conversion findings of earlier studies cited by the authors factors not stated) of Alba wheat grown in three (Mitchell and Hamilton 1929; Kon and Markuse locations ranged from 8.16% to 11.10%; Zanda 1931 ; Johnson and Palmer 1934). wheat at one location contained 9.08%. Petkuzer Sure ( 1954) compared the nutritive values of the rye at two locations contained 8.07% and 8.36% proteins (nitrogen conversion factors are not and Court des Flandres rye at two locations 7.82% and 8.80%. The mean lysine and threonine contents (Stein and Moore 1951) expressed as TABLE 58. Growth promoting values of wheat and percent total nitrogen were, in the wheats 2.22-and rye fed as sole source of protein in the diet for rats (data from Jones et al. 1948). 2.30 respectively and in the ryes 3.90 and 2.34. Janicki and Kowalczyk ( 1965) examined the Average protein content and amino acid composition (by Average gain/g Average autoanalyzer, tryptophan, Horn and Jones 1945), weight protein food of five varieties of wheat and three varieties of rye gains consumed consumed grown in the same areas of Poland and in the (g) (g) (g) same year. The results are presented in Table 59. Applying the method of Block and Mitchell 4. 5% protein in diet ( 1946) and comparing the biological value of egg protein with that of wheat, the authors found the Rye 35±2.40 2.26 337 limiting amino acids of all the wheat varieties to Wheat, hard 18±0.94 1. 72 234 be methionine and lysine. The limiting amino Wheat, soft 20± 1.32 1. 74 216 acids of rye were methionine and isoleucine. Rye 7. 5% protein in diet protein has a higher biological value than wheat protein mainly because it contains more lysine Rye 60±2.29 2.16 381 and methionine, but wheat has a higher protein Wheat, hard 43±2.11 I. 55 362 content than rye. Janicki and Kowalczyk cite Wheat, soft 28± 1.68 I. 21 326 Trzebska-Jeska and Morkowska-Gluzinska ( 1963) as having found in their work that tryptophan and 9. 5% protein in diet methionine were the limiting amino acids in rye. The latter however did not examine isoleucine and Rye 74±2.44 1.83 434 Janicki and Kowalczyk found higher values for Wheat, hard 68±2.65 1.60 444 Wheat, soft 66± 2. 35 1.68 414 tryptophan than did Trzebska-Jeska and Mor­ kowska-Gluzinska ( 1963). HULSE/LAING: PROTEINS OF TRITICALE 89

TABLE 59. Amino acid hydrolysates of Polish wheat and rye varieties (in grams/16 g N) (data from Janicki and Kowalczyk 1965).

Wheat Rye

Dan- kowskie Lagiew- Nagra- Zdzis- Przodow- selec- Wlosza- Rogo- nicka 52 Odin dowicka lawka nica ti on nowskie linskie

Protein N x 5.7 ~ 0 11.62 12.45 13.49 14.0 14.33 8.46 9.57 9.79 Alanine 3.83 3.71 3.49 3.36 3.21 4.80 4. 35 4.25 Arginine 4.66 4.89 4.90 4.52 4.17 5.59 5.55 Aspartic acid 5.41 5.39 5 .14 5.20 5.24 7 .13 6.82 7.46 Cystine (Cysteine and cysteic acid) I .43 l.46 I. 53 I. 55 Phenylalanine 3.71 3.74 4.12 3.99 4.10 2.71 3.68 3.34 Glycine 4.02 4.00 3.95 3.85 3.77 4.76 4. 36 4.25 Glutamic acid 27.0 27.7 28.0 28.2 28.0 20.9 22.3 21. 8 Histidine 2.29 2.39 2.33 2.54 2.25 2.46 Isoleucine 3.55 3.36 2.98 4.09 3.23 3.34 3.58 3.24 Leucine 6.73 6.51 6.30 6. 13 6.35 6.09 6.40 5.85 Lysine 3.02 2.87 2.80 2.79 2.66 4.37 3.94 4.09 Methionine (and methionine sulphoxide) l.46 I. 61 I. 56 l.44 l.41 I. 79 I. 76 I. 73 Pro line 8.66 9.55 8.58 10.3 IO. I 7.96 9. 17 8.66 Serine 3.94 4.00 4.02 3.93 3.71 4.45 3.68 4.01 Threonine 2.84 2.67 2.56 2.60 2.46 3.60 2.99 3.07 Tyrosine l.43 I. 77 I. 84 2.00 2.07 l.41 l.43 Tryptophan I. 11 I. 13 1.14 l.04 l.06 l.00 0.95 Valine 4.19 4.37 4.00 4.09 4.05 4. 50 4.53 4.39

Kalmykov (1968) used column exchange (BV) was determined using the dog as experi­ chromatography (Moore et al. 1958) to compare mental animal. Under the conditions of the the amino acid composition of Vyatka-2 rye and experiment, the BV of the rye was 89.5% and of Bezostaia-1 wheat. The results of these determina­ the wheat 76%. In Kalmykov's view the bio­ tions are given in Table 60. The biological value logical tests were in accord with the results of the chemical determinations of amino acid content. TABLE 60. Amino acid composition of Vyatka-2 rye Eggum (1970) examined the true digestibility and Bezostaia-1 wheat in grams/100 g protein (data of a number offeedstuffs, including wheat and rye, from Kalmykov 1968). and of amino acids contained in them, for baby pigs and rats. The results are presented in Table Rye Wheat 61 and indicate that amino acids from the same protein source can be digested differently, so Threonine 3.03 3. IO making it difficult to estimate the availability of Valine 4.64 3.90 any particular amino acid from the digestibility Cystine 2.8 2.7 Methionine I. 8 I. 8 of the total nitrogen. Isoleucine 3.34 3.72 Leucine 5.33 6.35 Tyrosine 2.2 2.3 Environmental and Agronomic Influences Phenylalanine 4.6 4.7 Pessi et al. ( 1971) studied the effect of fertiliza­ Lysine 4. 54 2.9 Tryptophan 0.87 I. 03 tion technique on the protein content of cereals grown in Finland in 1969-70. The fertilizer used 90 MONOGRAPH IDRC-02le

TABLE 61. True digestibility (TD)" for pigs and rats Calcium ammonium nitrate was found to be of total nitrogen (N) and the single amino acid for more effective than urea in improving grain and casein, wheat and rye (data from Eggum 1970). protein yield of Svenno and Apu spring wheats. Placement fertilization was more effective than Casein Wheat Rye broadcasting fertilization in increasing grain yield of spring wheat and total protein yield even though TD" for total N (pigs) 99.4 91. 8 80.9 the former also resulted in slightly lower protein TD" for total N (rats) IOI. I 89. 6 77.0 To• for: content in the grain. Cystine 100.1 93.9 86.3 Fertilization of winter wheat and rye with Aspartic acid 98.9 87. I 76.9 nitrogen in the spring improved protein content Methionine 100.3 88.6 73.9 without impairing yield, when compared to Threonine 98.8 88.4 74.4 fertilization in early winter. Additional nitrogen Serine 100.9 96.5 88. I fertilization at the heading stage increased the Glutamic acid 99.6 97.5 92.2 protein content. Glycine 97.7 89.7 78.2 Janicki et al. (1972) reported on the total Alanine 97.9 86.5 73.3 Kjeldahl nitrogen and amino acid composition Valine 99. I 90.8 78.0 (by autoanalyzer, excluding tryptophan) of five Isoleucine 99. I 90.3 73.7 Leucine 99.3 93.2 81 .2 Polish wheat flours, four of which were named Tyrosine 98.7 92.5 79. I varieties, and one rye flour designated only by Phenylalanine 98.8 92.0 79.8 a code number. On the basis of these, and un­ Lysine 99.7 84.1 72.2 published results, the authors suggest that a more Histidine 99.7 95.9 88.8 realistic nitrogen to protein conversion factor for Arginine 99.2 95.0 88.5 wheat flour is 5.61 instead of 5. 7 and for rye flour 5.67 instead of6.25 as is given in the Polish Official •True digestibility is not defined in the paper but is Standard. The protein contents of the flours generally regarded as the proportion of food nitrogen using both factors are given with the lysine, that is absorbed (National Academy of Sciences­ National Research Council 1963). methionine and threonine content in Table 62.

for winter wheat and rye was calcium ammonium Rye nitrate (26% N) and for spring wheat NPK Genetic and Varietal Effects fertilizer. The winter wheat varieties were Elo, Jyva, Linna, Nisu and Vakha and the rye varieties Trzebska-Jeske and Morkowska-Gluzinska were Ensi, Pekka, Toivo, Visa and Voima. (1963) conducted a survey of33 samples of winter Differences in yield were found between varieties rye harvested in 15 provinces of Poland in 1959. and with and without chlorcholine chloride (CCC) Of the 11 varieties grown, Ludowe and Wlosza­ treatment, but no statistically significant differences nowskie were grown in nine provinces, Pulawskie in grain crop protein were found which could be Wczesne and Dankowskie-Selekcyjne in four attributed to variety, fertilization or CCC treat­ provinces, and Zeelandskie, Universalne, Roga­ ment. linskie, Wierzbienskie, Mikulickie, Kazimierskie The six varieties of spring wheats are not named. and Wielkopolskie in one province each. As the level offertilization increased, grain protein No significant differences were found in the increased but CCC treatment had no effect nor mean Kjeldahl nitrogen values for the different were there statistically significant differences be­ varieties but considerable variations in nitrogen tween varieties. content and in weight of I 000 grains were observed The grain of one variety of spring wheat, within the same variety. For example, the mean J0-04558 grown in the two years 1968-70 was nitrogen contents of the widely grown varieties also analyzed for amino acid composition. were: Ludowe 1.44% ranging from 1.30 to 1.67% (Methods of determining protein and amino acid and Wloszanowskie 1.39% ranging from 1.17 to composition not stated.) The percentage of pro line 1.58%. Lysine, leucine, tryptophan, arginine and and glutamic acid increased and the percentage methionine (microbiological assay) as percent of lysine decreased with increased fertilization. total nitrogen also varied more within varieties HULSE/LAING: PROTEINS OF TRITICALE 91

TABLE 62. Protein. ash. lysine. methionine and threonine contents of five Polish wheat flours and one rye flour (data from Janicki et al. 1972).

Wheat flours Rye flour Luksusowa Wroclawska Krupczatka Tortowa Code No. Code No. Code No. Code No. Code No. Code No. 750 500 950 500 450 800

Moisture. ~~ 10.5 11. 3 10.7 IO. I 10.4 9.0 Ash 0.841 0.553 0.926 0.482 0.485 0.705 0 Kjeldahl Nitrogen. 0 I. 97 I. 79 2.16 1.48 1.64 1.27 0 Protein N x 5.7. 0 11. 23 10.20 12.06 8.46 9.35 7.96 0 Protein N x 5.61. 0 (wheat) 11.05 10.05 11. 87 8.33 9.20 0 N x 5.67. 0 (rye) 7.22 Amino acids as percent of total amino acids Lysine 2.60 2.49 2.68 2.40 2.57 3.54 Methionine 1.46 I. 55 I. 31 1.60 1.29 1.44 Threonine 2.95 2.83 2.98 2.81 2.75 3.77

TABLE 63. Origin. moisture. and (Kjeldahl) nitrogen. and nitrogen efficiency ratio of four rye flours fed to rats (data Kihlberg and Ericson 1964).

B c D 50% 20% 70% Russian Russian Russian A 50% 80% 30% Russian Swedish Swedish Argentinian

Moisture.% 12.8 13.9 13.2 12.0 Nitrogen. % of fresh weight of flour 2.03 I. 75 1.47 I. 79 Rat feeding trials Average weight gain g/day 1.63±0.10 I. 7 ±0.07 2.09±0.10 1.55±0.08 Nitrogen efficiency ratio g weight gain/g nitrogen eaten 12.25±0.29 13. 32±0. 31 14. 66 ±0.47 11.51±0.38 than did the mean values. Tryptophan, methionine which was, in grams/ 16 g N, for (A) 4.13, (B) 3.41, and lysine were inversely correlated with total (C) 3.16 and (D) 2.88. The rye flours, 91 % extrac­ nitrogen, though total content of the amino acids tion rate, supplied all the protein and carbo­ increased with the increase in protein content. hydrate in the diets for young male albino rats, Kihlberg and Ericson (1964) compared four eight per group, fed for 28 days, the diets providing rye flours designated (A) Russian, (B) 50% 1.34% nitrogen. The average weight gain and Russian, 50% Swedish, (C) 20% Russian, 80% nitrogen efficiency ratios are also given in Table 63. Swedish, (D) 70% Russian, 30% Argentinian, for Lysine was found to be the first and threonine the amino acid composition (ion exchange chroma­ second, most limiting amino acid. No single amino tography, except for tryptophan and cystine, acid or pair of acids increased the protein value of assayed microbiologically), and for biological rye flour supplemented with lysine and threonine. quality in rat feeding trials. The moisture content Zhebrak and Kolchin (1968) found little dif­ and Kjeldahl nitrogen are shown in Table 63. ference in the amino acid content (by paper The amino acid composition of the four rye flour chromatography), expressed as percent rye pro­ mixtures were essentially similar except for lysine tein, in the grain of diploid and tetraploid rye of 92 MONOGRAPH IDRC-02le

TABLE 64. Moisture, nitrogen, protein, 1000-grain weight, lysine and threonine contents of spring and winter rye grain (data from Boronoeva and Kazakov 1969).

Spring rye Winter rye

2 3 4 5

Moisture content, % 8.6 9.0 8.4 8.6 10.4 Weight of 1000 grains, g 33.9 29.7 24.9 16. l 16.2 Nitrogen % dry weight 2.69 2.30 2.65 1.90 2.36 Protein (N x 5. 7), % 15.30 13 .11 15. IO 10.80 13.45 Amino acids as g amino acid per I 00 g protein Lysine 2.56 2.47 2.65 2.38 2.38 Threonine 4.80 4. 35 4.05 3.68 4.20 the Petkusskaya variety. The 1000-grain weights (2) Six different varieties grown m the Minsk were, respectively, 26.66 g and 57.30 g. district, Boronoeva and Kazakov (1969) determined by (3) Four varieties grown in three places. autoanalyzer the amino acid composition of (I) Total protein content (N x 6.25) ranged from Onokhoisk spring rye harvested in 1964 at the 8.6% to 14.4%. The means were: Group I, 12.3% Buryat Agricultural Research Station, (2) Onok­ (range 10.6 to 13.6%). Group 2, 11.5% (range hoisk spring rye harvested in 1965 at the same 9.9 to 13.3%). and Group 3, 12. I% (range 8.6 to station, (3) Onokhoisk spring rye harvested in 1965 14.4%). There was relatively little difference in in the Barguz region of the Buryat ASSR, (4) the amino acid content as percent rye protein in Udinsk winter rye harvested in 1964 at the individual rye varieties, e.g. lysine ranged from Buryat Agricultural Research Station, (5) Winter 3.1 to 4.2; the means were: for Group I, 3.9 (range rye harvested in 1966 in the Orenburg region. 3.6 to 4.1 ), for Group 2, 3.9 (range 3.1 to 4.2), Table 64 shows the nitrogen, moisture content, Group 3, 3.6 (range 3.4 to 3.8). protein, 1000-grain weight, lysine and threonine Schneider and Lantzsch (1971) reported on the contents of the spring and winter ryes. There were apparent digestibility of three winter ryes and two no substantial differences in the amino acid com­ summer ryes in feeding trials with sheep, four per position between winter rye and spring rye. group and pigs, three per group. The ryes differed Minja (1969) reported briefly that when the rye in their crude protein content as is shown below: varieties Antelope, Dakold, Prolific 6201 and 6203 were, (I) without treatment, (2) autoclaved, (3) Crude protein (N x 6.25) solvent extracted, each fed to mice, autoclaving (% dry matter) had a detrimental effect; solvent extraction had a Karlshuder winter rye 13.62 beneficial effect on 6201 and 6203. Rations con­ Karlshuder summer rye 14.44 taining Dakold and 6204 were equivalent in Petkuser winter rye, short straw 9.29 apparent biological value to the casein control Petkuser winter rye, normal straw 9.14 diet containing 12% crude protein but the treat­ Petkuser summer rye 10.17 ment, if any, that the rye received is not stated. In spite of these differences in crude protein con­ tent, the differences in the apparent digestibility of the ryes were not significant with either sheep Environmental and Agronomic Influences or pigs. Golenkov and Gilzin ( 1971) determined (by Somin (1970) determined the amino acid com­ autoanalyzer) the amino acid content of22 samples position (ion exchange chromatography) of 15 of basic varieties of rye from the 1967 Russian varieties of rye grown in the USSR under a variety harvest and 19 from the 1968 harvest. of environments. The varieties were divided into About one-third of the amino acid content as three groups: percent protein was in the form of glutamic acid (I) Vyatka 2, grown in five different areas, (from 21.74 to 35.15%) and proline (8.45 to HULSE/LAING· PROTEINS OF TRITICALE 93

12.99%). In all samples examined there was a high respectively, (I) 4.04, (2) 3.92, (3) 3.65, (4) 4.09, content of lysine (3.00 to 5.30%), of threonine (5) 3.99 and threonine (I) 3.24, (2) 3.45, (3) 3.43, (2.82 to 3.91%) and of valine (3.72 to 7.25%). (4) 3.45, (5) 3.24. Statistical analysis suggests the variations within Fernandez et al. (1972) has reported that when amino acids are real and significant and not laying hens were fed a diet containing a high level entirely attributable to analytical errors. (80%) of rye (of unstated origin), there was a sharp The extent of these variations, however, differs drop in the rate of egg production in the first three considerably from one year to another. In almost weeks followed by an increase in egg production to every case, the variations were smaller in 1967 near normal level. The drop was not prevented by than in 1968, which may be explained by the fact supplementation with penicillin. that the environmental conditions were less Chlorcholine chloride (CCC) treatment, which variable in 1967 than in 1968. Results quoted also has been shown to reduce the height of rye show that different varieties grown in the same (Primost et al. 1967) was applied, with nitrogen place, and one variety, Kharkovskaya 60, grown fertilization, to Schlagher alt and Kefermarkter in different regions, were significantly different in varieties of rye at Linz/Donau, Austria. Both their amino acid contents. these varieties had responded to treatment with The content of lysine in the total protein of the CCC (Bayzer and Mayr 1967). varieties Kharkovskaya 55, 60 and 194 grown at CCC-treated samples of the same (Kjeldahl) the Kharkov Genetic Institute in 1967 varied only nitrogen content as untreated samples had approxi­ slightly, but in 1968 the variations exceeded the mately the same amino acid composition (method error of determination by three times. A similar of Moore et al. 1958; tryptophan by micro­ picture emerged for the lysine content of Sara­ biological assay) and there was therefore no in­ tovskaya, Krupnozernaya and Volzhanka grown crease or decrease in amino acid composition as a in the Saratovskii region. Lysine (percent grain) direct result of CCC-treatment. Primost et al. from the varieties Kharkovskaya 60, Hybrid 2 state that the higher the nitrogen content in rye, and Vyatka Moskovskaya grown in Stupino the more glutamic acid and proline and the less (Moscow district) was dependent neither upon lysine, arginine and aspartic acid. Differences in harvest season nor variety. the variety, location and fertilization of rye affect The amino acid content of Vyatka rye did not the amino acid composition only in so far as they appear to be significantly affected when grown affect the nitrogen level in the grain. The same following: (I) fallow, (2) peas, (3) maize, (4) vetch effect was produced by CCC-treatment during the and oats, (5) barley, e.g. lysine content was, vegetative period. Kernels of triticale infected with ergot ( Photo : £. N. Larter J Chapter 4

NUTRITIONAL INHIBITORS AND Toxic FACTORS

Introduction weight increase of cattle, sheep, pigs, poultry, horses and rabbits, has been recognized for many The nutritional value of a cereal may be im­ years and was investigated by Wieringa ( 1967) in paired by the presence of naturally-occurring the Netherlands. factors which interfere with the digestibility, and/or In experiments using Petkuser winter rye, availability to human and/or other animals of one harvested in 1961, fed to weanling male white or more of its nutrient components. The distinc­ Wistar rats, he identified the growth-inhibiting tion between a nutritional inhibitor and a toxic substances as a mixture of 5-n-alkyl resorcinols substance is indeed fine. The word ""toxic" is with odd numbered side-chains of I 5-23C atoms, defined as "poisoned or imbued with poison" and together with smaller amounts of 5-alkenyl is derived from the Greek name for the poison resorcinols. used for smearing the tips of arrows (w~1Koa, The same group of compounds had been found equals or pertaining to the bow). A poisonous (or in wheat by Wenckert et al. (1964). No difference toxic) substance is one which when introduced or could be found by Wieringa ( 196 7) in growth­ absorbed by a living organism destroys life or inhibiting effect between rye and wheat resorcinols, injures health. Consequently a nutritional in­ in experiments in which the rye used contained hibitor, if its inhibiting effect is sufficiently potent about twice the level of resorcinol as the wheat. to cause serious malnutrition, might properly be He found great variation in alkyl resorcinol con­ described as "toxic". tent within a single sample of rye. He also found The nutritional inhibitors reported in triticale, that the percentage of crude protein correlated wheat or rye, fall into two broad categories: positively with 1000-kernel weight, and that the (A) Intrinsic substances which occur naturally heavier kernels contained relatively less resorcinols in the plant. These include resorcinols, phytates, than the lighter kernels. It is possible, at least in certain enzyme inhibitors, and others as yet part, that variation in resorcinol content was unidentified. The presence of nutritional inhibitors attributable to variations in grain composition, is detected by a lower than anticipated rate of the lighter grains probably displaying a higher weight increase in the animal consuming the food bran to endosperm ratio than the heavier grains. in relation to the quantity and quality of the Chromatographic examination of the rye kernels nutrients ingested. established that the resorcinols are localized in the (B) Substances produced by infection with pericarp and do not occur in the endosperm or microorganisms. Many of these, including ergot, germ. Consequently scientists who encounter aflatoxins and other mycotoxins are, as their apparent varietal differences in inhibitor content name implies, highly toxic. should ensure that the differences are truly genetic in origin and not in fact attributable to differences Naturally-Occurring Inhibitors in end_osperm size, varying degrees of shrivelling or grain malformation. Resorcinols Wieringa also established that older rats, with The observed effect of rye, when fed in large a body weight of about 100 g became accustomed amounts, in reducing the food intake and rate of to a diet including the growth-inhibiting factor

95 96 MONOGRAPH IDRC-02Ie whereas weanling rats of approximately 40 g resorcinols, the content of alkyl resorcinols in body weight did not. triticales being lower than in rye but higher than In experiments with 20 farrows of 'VNL' in wheat. piglets basic rations were fed to which were Munck ( 1972) also reported on the resorcinol added (A) 50% barley meal (control), (B) 50% rye content of seven commercial milling fractions meal, variety D 1964, (C) 50% rye meal, Petkuser, from a Swedish wheat (see Table 88 in Chapter 5). 1964, (0) 50% rye meal, Petkuser, 1965, and (E) There was high positive correlation (r = 0.98) 49.15% barley meal with 0.85% rye oil. The between crude fibre and resorcinol content, again average body weight of each group of 10 animals, indicating the association of resorcinol with cer­ half male and half female, was 16.2 kg. During the tain bran components. experiment diarrhoea occurred and six animals Tests in England, employing weanling rats have died, two from the control group. No symptoms of confirmed the presence in wheat bran oil of an over-feeding with rye were observed but the unidentified substance which depressed both growth results for all groups were lower than appetite and growth rate (Flour Milling and expected. Baking Research Association 1971 ). Most, though Taking groups (B), (C), (0), (E) together, the not all, of the reduction in growth rate was due to average growth was 11 to 12% lower than that of reduced food intake. The smallness of the residual the control group. This difference was highly inhibitory effect indicates either that the amount of significant (P = 0.01) though differences between material in bran, or its potency, is small. each test group separately and the control were Wieringa ( 1967-68) has further reported that, in not significant (P = 0.10). the Netherlands, more of the growth-inhibiting Munck ( 1969) reported that at Svalof, in Sweden, substance is derived from the by-products of a preliminary screening of 5-alkyl resorcinols in wheat milling than from rye because a much larger rye showed great variation with genetic and quantity of by-products incorporated into animal environmental conditions, ranging from 0.12 to feeds is derived from wheat than rye. 0.22% content of the seed (moisture content about The occurrence of phenolic compounds in the 10%). In winter rye of low protein content 5-alkyl young leaves of one hexaploid and three octaploid resorcinols were present in higher concentrations triticale lines and of their wheat and rye parents than in winter rye of higher protein content. Munck was studied by Dedio et al. ( 1969). Methanol-HCI also reported that spring varieties of rye showed extracts, analyzed by two-dimensional thin-layer no such correlation but gave no further details. chromatography, showed a compound present in In wheat, lower amounts of the same group of large amounts in rye to be transmitted to the compounds were found. triticale. A later report (Munck 1972) on the alkyl Villegas ( 1972) gives the resorcinol content of 13 resorcinols in rye, wheat and octaploid triticale strains of triticale as ranging from 0.070 to grown in Sweden gives the findings presented in 0.092% of sample. Table 65. The octaploid triticales resemble wheat Although the growth-inhibiting effect of 5-alkyl more closely than rye in their content of alkyl resorcinols in some animals has been demon­ strated, their significance in human diets has not TABLE 65. Alkyl resorcinol "units" (range in paren­ yet been determined. Munck ( 1969) found that theses) in rye, wheat and (octaploid) triticale (data from 30% of the 5-alkyl resorcinols present disappeared Munck 1972). during the baking of soft rye bread. Zillman (1973, unpublished data) evaluated the Alkyl resorcinols effect of resorcinol on growth rate of mice by add­ Cereal No. of varieties ("units") ing, in predetermined amounts, ( 1) the acetone­ extracted resorcinol compound from rye to diets Rye 15 161 of triticale, rye, and wheat prepared in the form (129-192) of wafers, and (2) by physically mixing predeter­ Wheat 18 69 mined quantities of bran (which contained the (56--79) resorcinol fraction) to the flour of the grains. Triticale (octaploid) 19 97 (78-124) The results showed that the resorcinol content of grain per se is not detrimental either to growth rate HULSE/LAING. PROTEINS OF TRITICALE 97 or feed intake of mice. Contrary to certain reports practical purposes. all of the calcium present is in the literature. the feeding value of both triticale rendered unavailable in I 00% extraction flour. and rye was superior to wheat and Zillman con­ Consequently in some countries during wartime cludes that in previous studies in which rye and legal requirements were introduced to demand triticale were found to result in poor growth. the calcium supplementation of high extraction flours. effects could most likely be attributed to the Hay ( l 942)cited by Kent-Jones and Amos( 1967) presence of ergot. reported that in commercial wheat products. phytic acid content runs parallel to fibre content. Phytic Acid Phytic acid and the phytates may be decomposed Phytic acid (inositol-hexaphosphoric acid) and by the enzyme phytase to produce inositol and its derived salts. the phytates. occur in the seed phosphoric acid. Wheat and ryeare rich in phytase. coats of many cereal grains. In cereals. phytic which is unequally distributed through the grain acid represents 35 to 97~~ of the total phosphorus in somewhat the same manner as phytic acid. The (Gontzea and Sutzescu 1968) and is generally phytase of rye is more active than that of wheat regarded as a poor source of phosphorus to (Gontzea and Sutzecu 1968). The extent to which monogastric animals. It is not equally distributed. the phytic acid content of wheat and rye may be the pericarp and adherent aleurone layer, together harmful in a diet will depend on the conditions with the germ. containing proportionally IO to 20 under which the cereal has been processed for times the level in the endosperm. The importance consumption. of phytic acid in nutrition depends upon its Kent-Jones and Amos ( 1967) cite Pringle and property of forming insoluble or nearly insoluble Moran ( 1942) who reported on the effect of yeast compounds with mineral elements. including quantity and length of fermentation time upon the calcium. iron. magnesium and zinc. the resultant destruction ofphytic acid in bread made from 85% phytates being excreted in the faeces. Consequently extraction wheat flour. This effect is shown in dietary calcium. iron. etc. can be rendered unavail­ Table 67. The quantity of yeast had only a small able when combined with phytic acid. Diets high effect but the longer the fermentation. i.e. the in phytic acid and poor in calcium. iron and zinc. longer the period during which the phytase could produced mineral deficiency symptoms in experi­ act on the phytic acid. the greater the reduction in mental animals and in children (Gontzea and phytic acid. Because of the distribution of phytic Sutzescu 1968). acid and phytase. and the effect of baking tempera­ The quantity ofphytic acid found in the products tures. white bread has a lower content than whole of milling will therefore depend to a large extent wheat bread. and rye bread has a lower content on the extraction rate. Urie and Hulse ( 1952) quote than wheaten bread of equivalent extraction rate. data given in Table 66 to illustrate the increase in Table 68 indicates the position (Gontzea and phytic acid phosphorus with increase in wheat Sutzecu 1968 ). flour extraction rate. They state that the available Gontzea and Sutzecu (I 968) also state that (soluble) calcium present in 72% extraction flour cereals prepared by heating in boiling water or is of the order of I 0 mg calcium per I 00 g flour. Because of the increase in phytate phosphorus. TABLE 67. Effect of fermentation time and yeast barely I mg per I 00 g available calcium is present quantity on destruction of phytic acid during f ermenta­ in I 00~~ extraction (wholemeal) flour. For all tion and baking 85% extraction wheat flour (data from Pringle and Moran l 942). TABLE 66. Increase in phytic acid phosphorus with increase in wheat flour extraction rate (data from Proportion of Urie and Hulse 1952). Length of Proportion of phytic acid fermentation yeast destroyed Extraction rate Phytate phosphorus (hours) (%) (%) ('%,) (mg/ JOO gflour) 3 59 72 50 5 64 1 85 130 5 2 61. 5 100 300 8 76 98 MONOGRAPH IDRC-02\e

TABLE 68. Phosphorus and phytic phosphorus in wheat and wheat products, and rye and rye products (ranges, where available, in parentheses). (Gontzea and Sutzecu 1968 citing various references which are not specifically identified against the items).

Total Phytic phosphorus phosphorus (P) (mg/JOO g) (mg/JOO g) ('\ of"total P)

Bread wheat 320-360 170-280 47-86 Rye 342 247 73

Wheat flour 100/0 430 307 71 (356-567) (239-385) Bread from wheat flour I 00% 415 210 51 (330-485) (185-260) Rye flour 100% 328 168 64 (226-401) (120-276) Rye flour 95% 302 169 56 (219-386) (110-236) Bread from rye flour 95% 286 32 II (235-342) (0-68) Wheat flour 70% 218 125 57 (169-285) (66-194) Bread from wheat flour 70% 226 46 20 ( 178-276) (10-102) Rye flour 60% 155 62 40 (136-175) (45-82) Bread from rye flour 60% 143 0 0 (116-156) Wheat bran 1210-1609 1170-1439 89-97

milk may be higher in phytic acid than those pre­ TABLE 69. Phytic acid phosphorus in bread ingre­ pared by baking since the boiling fluid inactivates dients and bread made from white flour with additions the phytase before it can decompose the phytic of wheat protein concentrate (WPC) (data from acid. Autoclaving or prolonged boiling will, to Ranhotra et al. 1971 ). some extent, decompose phytic acid even in the absence of phytase, the degree of decomposition Phytic acid phosphorus, being greater under acid conditions. moisture free sample Ranhotra et al. (I 971) determined the phytic (mg/JOOg) acid phosphorus in the ingredients and in the resultant baked bread when progressive incre­ Flour + 0% WPC Bread ingredients 13.6 ments of wheat protein concentrate (WPC) from Bread 0.0 a commercial mill replaced equivalent amounts of the flour in the blend. The white flour, from hard Flour + 15% WPC red winter wheat, had a proximate analysis of Bread ingredients 76.8 protein (N x 5. 7) 11.83%, ether extract 1.28%, Bread 34.7 fibre 0.33%, ash 0.43% and moisture 11.07%. WPC, of protein 15.35%, ether extract 3.18%, Flour + 30% WPC Bread ingredients 130. 7 fibre 1.22/~, ash 1.97% and moisture 11.52% Bread 71.0 replaced 15%, 30% and 45% of the flour. Phytic acid was estimated by the method of Anderson Flour + 45% WPC (I 963). The effect of these substitutions, and of Bread ingredients 190.4 the baking process is shown in Table 69. The results Bread 132. 2 show the increase in phytic acid with increase in HULSE,LAING: PROTEINS OF TRITICALE 99

WPC and the reduction which occurs during Enzyme Inhibitors baking. Trypsin is one of the proteolytic enzyme systems Ranhotra ( 1972) has further reported that when present in the internal secretions and essential to WPC progressively replaced, up to I 00%. hard the efficient digestion of ingested protein. The red winter wheat flour in blends baked into literature on protease inhibitors was reviewed by small loaves by a sponge-dough procedure. the Liener and Kakade ( 1969). Trypsin inhibitors percentage of phytic acid hydrolyzed was inversely have been identified in wheat and rye but their related to the amount initially present. All the importance in practical nutritional terms remains phytic acid phosphorus was hydrolyzed during obscure. baking of the 100\ wheat flour blend. The amount Laporte and Tremolieres ( 1962) reported an hydrolyzed increased up to the 30° 0 level of WPC anti-trypsin factor in uncooked wheat and rye replacement; thereafter there was no increase in flour. In cooked flour (tubes plunged into boiling hydrolysis and the amount of phytic acid hydro­ water for 5 min. giving an effective temperature of lyzed was also inversely related to the amount 98cC for 4 min) no trypsin inhibition was found in present in the unbaked ingredients. The pH of the wheat flour but rye flour retained 80% of the breads baked from the blends increased progres­ inhibitory activity. Learmonth and Wood ( 1963) sively ( Ranhotra 1973) with WPC replacements also reported that cooking reduced the inhibitory from 5.20 (all wheat flour blend) to 5.75 (all WPC activity of wheat. bread) above the optimum for phytase activity in Milner and Carpenter ( 1969) determined a wheat of 5.1 to 5.3 (Peers 1953). Ranhotra ( 1973) trypsin inhibitor in an American hard winter wheat suggests that the progressive decrease in hydrolysis of approximately 17"/0 protein (N x 6.25). dry of phytic acid may be caused by increased inhibi­ weight basis. by two methods. (I) using haemo­ tion of phytase activity or by rephosphorylation globin as substrate. and (2) using benzoyl­ of partially hydrolyzed phytic acid. or by both. arginine p-nitroanilide. Measurable trypsin in­ Reinhold ( 1971) has reported on the differences hibition was recorded in extracts from the control, in phytic acid present in Iranian bread made untreated. wheat by both analytical methods but (a) in the village and (h) in the city. Village bread extracts from a sample of wheat steeped. as in the (tanok or lavosh) is made without fermentation. preparation of bulgur. for 70 min in water at 80cC In the city bakeries bazari and sangak bread are and then dried at a temperature below 40cC made by a 2- to 4-hour fermentation process showed no inhibitory activity. followed. for bazari. by baking on the roof of a Polanowski ( 1967) identified a trypsin inhibitor brick oven or. for sangak. on pebbles. for about in the aqueous extract of rye. It was located only 2 min. The wholemeal flours used in tanok and in the endosperm and not in the germ or seed coats sangak contain nearly all the original phytic acid which included the aleurone layer. Heating the of the wheat. the meal used for bazari contains extract at 75°C resulted in loss of inhibitory rather less. Samples of the three breads were activity, the inhibitor being completely inacti­ obtained over several months and examined for vated after heating for 25 min. phytic acid concentration by precipitation as Cho C. Tsen ( 1973. personal communication) ferric phytate in 0.6% HCI. The mean contents of writes: .. A trypsin inhibitor and a chymotrypsin phytate in milligrams per I 00 g dry bread were: inhibitor are extracted from triticale. wheat. and bazari bread. 36 samples. 301 (SD 96.9); sangak rye flours by means of a sodium acetate buffer at bread. 30 samples. 401 (SD 116.5) and tanok pH 3.8. The inhibitor content of triticale is inter­ bread. 96 samples. 630 (SD 140.0). mediate between wheat and rye, with the rye In both the villages and the city. flat bread exhibiting the highest trypsin inhibitor content provides at least half the caloric intake. It is sug­ and the wheat the highest chymotrypsin inhibitor gested that the higher phytate intake of the village content. The molecular weight of the trypsin population. who eat tanok bread. may account inhibitor as determined by molecular exclusion for the evidence of zinc deficiency in the village chromatography is 12-14,000. The trypsin in­ population. in spite of seemingly adequate zinc hibitor is thermally stable at I 00°C for one hour. intake. and also for the prevalence of rickets The molecular weight of the chymotrypsin inhi­ among village children, and the high incidence of bitor determined as above is 17-19.000. Thermal iron deficiency anaemia. inactivation occurs after I 0 minutes at 70°C. 100 MONOGRAPH IDRC-02le

Polyacrylamide gel disc electrophoresis of both improvement with penicillin, but the supplements, inhibitor fractions indicates the presence of several in combination, were not more effective than proteins." penicillin alone. Inactivation of the enzyme products by autoclaving destroyed the property Growth-Depressing Factors of chick growth improvement (McGinnis 1972). Experiments in which rye was used to replace Friend (1970) reported on experiments with other cereals at high levels in chick diets have male Wistar rats, averaging 167 g initial body shown that it has a low nutritional value for weight, fed ad libitum a basal diet in which 42% chicks of both sexes (MacAuliffe and McGinnis of the total diet was provided by (A) ground barley, 1971 ). Rye was treated (I) by adding an equal (B) ground rye, (C) barley flour plus rye bran, weight of water to the ground grain, spreading it (D) rye flour plus barley bran. Rats given diet (C) on trays and oven-drying at 70°C, and (2) by containing barley flour plus rye had the highest adding an equal weight of water, autoclaving for feed intake, gained most weight and retained more 15 min at 121°C and then oven-drying at 70°C. nitrogen than did rats fed the other diets. The least Water-treatment of the rye gave rise to a significant successful diet was (B) ground rye. Weight gains improvement in chick growth and feed efficiency. on diet (D) rye flour plus barley bran were better Alternatively the harmful effect of high levels of than the ground rye diet but less than on the other rye were largely overcome by addition of the two diets. The rye was ergot-free and it would seem antibiotics terramycin or procaine penicillin G that it is rye flour, rather than rye bran, which is but not by zinc bacitracin. MacAuliffe and associated with the lower feed consumption McGinnis suggest that the low value of rye is associated with diets containing rye grain. related to a component, possibly the high pentose Attia and Creek (1965) used male chicks in content, which stimulates the growth of adverse experiments to establish the nature of a growth microflora in the intestinal tract of the chick. depressant factor in raw wheat germ. Raw and Acetone extraction of rye, which would be autoclaved wheat germ were compared and the expected to remove any resorcinols present, did tests indicated the presence of a haemagglutinin not remove the growth-depressing effect of rye in factor in wheat germ which could be destroyed by chick diets in experiments described by McGinnis autoclaving. The work was carried out partly to (1972). Furthermore, the acetone extract of rye meet the criticisms of Parrish and Bolt ( 1963) who did not depress chick growth when added to a did not accept the presence of an anti-proteolytic diet containing wheat. factor in raw wheat germ reported by Creek and Water extraction of rye significantly improved Vasaitis (1962), and considered that the reduced its feeding value for chicks to a level almost equal weight gain then found was caused by a lower feed to, and not statistically significantly different consumption due to a paste forming on the beaks from, the improvement obtained by adding peni­ of the birds fed the raw wheat germ. cillin. The addition of penicillin to the water­ Cave et al. (1965) found that raw wheat germ extracted rye gave a further improvement. How­ meal when fed as the only source of protein to ever, adding the water extract of rye to a control chicks, produced poor growth and net protein diet based on maize did not depress chick growth. utilization. Mild acid treatment, followed by autoclaving, Olsen ( 1967) referring to the work described improved the nutritional value of rye in chick above on chicks, used male weanling rats to try diets, a further improvement being observed when and establish whether or not an antinutritive penicillin was added. Penicillin also improved the factor was present in raw wheat germ. The criteria untreated rye diet. The difference between the used were (a) amino acid composition, (b) digesti­ acid-treated rye and the untreated rye diets, when bility of the protein, (c) absorption of nitrogen and penicillin was added, was not statistically signi­ amino acids, and (d) effect of supplementation ficant. Alkali treatment of rye, followed by auto­ with amino acids on the utilization of the protein. claving, did not effect improvement (McGinnis The samples of wheat germ were (i) raw, (ii) toasted 1972). for 45 min at a product temperature of 121°C, Three commercial enzyme products, Rhozyme (iii) autoclaved, 15 lb. pressure (121°C) for 20, H39, Rhozyme CL and Lipase B, improved the 45 or 90 min. Olsen found no evidence of a utilization of rye by chicks comparably to the nutritional inhibitor for the rat in raw wheat germ HULSE/LAING: PROTEINS OF TRITICALE IOI and recommended that any heat treatment applied TABLE 70. Ergot infection in wheat and triticale to wheat germ should be as mild as possible since grown in two locations in North Dakota in 1971 (data excessive heating impaired the biological value of from Busch and Wilkins 1972). the protein. Per cent ergot (weight)

Toxic Substances Fargo Carrington Ergot Cereals 1971 1971 Ergot, which results from the infection of grain Wheats or grasses by the fungus, Claviceps purpurea, has Era 0.01 0.01 been recognized for a considerable time as a source Waldron 0.25 0.13 of damage to crops and of toxicity to livestock fed infected grain (Seaman 1971 ). The fungus prevents Triticales Rosner 0.53 seeds of the host plant from developing and re­ 0.06 6T A203 (FasGro 203) 0. 39 0. )8 places many kernels with hard, seed-like fungus 6T A204 (FasGro 204) 0.92 0.25 bodies known as sclerotia. 6TA209 0.86 0.47 The size and shape of the sclerotia vary with the Trailblazer (a reselection host plant. The sclerotia in rye are often horn­ of6TA209) 0.62 0.22 shaped and several times larger than the kernels. Grains in which some sterility may occur, such as triticale, and cross-pollinated grains such as rye, Wide variations in the tolerance of livestock to are more likely to be infected because their florets the ergot toxin have been recorded. In Canada, are open longer than those of fertile, self-pol­ limits are imposed on ergot in grain and Seaman linated hosts. Among cultivated crops, rye and (1971) states that feed containing 0.1 % ergot triticale are more often infected than wheat, and should be regarded as dangerous. among the wheats T. durum is more often affected Bragg et al. (1970) however, investigating the than T. aestivum. effect of triticale ergot on the performance and Treatment of the seed with fungicide does not survival of broiler chicks found they could tolerate appear to control ergot. Zillinsky and Borlaug up to 0.8% ergot in the diet. They recommended, (197lb) state, however, that Larter has reported however, that commercial feed should not con­ the wheat variety Kenya Farmer as being resistant tain more than 0.4%. to ergot. Crosses between Kenya Farmer and Two feeding trials were carried out simulta­ triticale varieties have been made at CIANO in neously with 330 commercial broiler chicks dis­ 1970 in an attempt to transfer resistance to ergot tributed at random into 33 experimental pens of from Kenya Farmer to triticales. Crosses were I 0 birds each at one day of age. In the first trial also made between Kenya Farmer and several eight levels of triticale ergot (0, 0.2, 0.4, 0.8, 1.6, varieties of rye to produce new octaploid triticales. 3.2, 6.4 and 12.8%) were supplied in a basal ground E. N. Larter (1973, personal communication) wheat ration, at the expense of wheat. Dietary informed the authors that similar attempts are treatments were arranged in a randomized block being made at the University of Manitoba to design. In the second trial, three dietary treatments build ergot resistance into triticale by crossing were prepared with a ground triticale basal ration with Kenya Farmer and other resistant wheat (the triticale at the same weight for weight level as varieties. The results will not be available until in the wheat ration). Ergot was supplied at two late in 197 4. levels (0 and 1.6%) at the expense of the triticale. The problem of ergot infection of triticale is Average weight gain, feed: gain ratio (F/G) and illustrated in the findings of Busch and Wilkins percent mortality were calculated at the end of 28 (1972) for five triticales and two wheats grown at days and 56 days of feeding the experimental diets. Fargo and at Carrington, North Dakota in 1971. In the first trial results at 28 days of age showed The figures are given in Table 70. Ergot infection no significant difference in weight gain between was, in general, higher in triticale than in wheat, chicks fed 0.4%, 0.8% ergot or the control diet. and, again in general, higher at Fargo than at Maximum weight gain was obtained with 0.2% Carrington. ergot which was significantly higher than 0.8% 102 MONOGRAPH IDRC-021e dietary ergot. A very severe depression in weight TABLE 71. Effect on dry matter intake, weight gain gain was shown in chicks fed 1.6% dietary ergot and feed efficiency of dairy calves fed ergot-containing and the depression continued as the dietary level diets (data from Ingalls and Phillips 1971). increased until growth practically ceased at 12.8/0 dietary ergot. The F/G ratio was not adversely Dry affected at ergot levels of0.2, 0.4, and 0.8%; there matter Weight Feed was a small but consistent increase with increasing intake gain effi- (kg) (kg/day) ciency levels, the result of a voluntary restriction of feed intake. Dietary levels of 1.6% and above showed Barley 3.39 0.97 3.50 very poor F/G ratios. Barley+ 0.07% ergot 3 .10 0.84 3.71 Chicks fed ergot diets from 29 to 56 days of age Triticale + 0.07 - showed the same growth pattern observed during 0.1% ergot 2.89 0.85 3.42 the first 28 days. There was no significant growth Barley+ 0.14% ergot 3.20 0.88 3.68 depression until the 1.6% level of dietary ergot was Barley+ 0.28% ergot 2.61 0.72 3.20 reached. The F/G ratio was not severely affected even at the 1.6% level fed to 56 days of age. Ergot levels of 3.2% and above however led to signs of McCloy et al. (1971) cite E. W. Stringham toxicity and by 48 days of age all the chicks in the (1968) and H. H. Nicholson (in a personal com­ 3.2% ergot group had died and by 56 days the munication) who observed reduced performance mortality in the 1.6% group was 23%. in cattle fed triticale, compared to barley and In the second trial weight gain and F/G ratio maize, but who also noted relatively high, but were equal for chicks fed ergot-free wheat and unstated, levels of ergot in the triticale used. triticale diets. The addition of 1.6% ergot to Harrold et al. (1971) comparing triticale from either wheat or triticale diets depressed growth. two different crop years as a feed for growing­ Pelleting the triticale feed increased the growth finishing swine used grain which contained ergot depression significantly compared to the wheat, for part of the trials but the level of contamination triticale, or 1.6% ergot and triticale diets in mash is not stated. The results from cleaned 1968 crop form. were however inferior to those from uncleaned In experiments conducted by Ingalls and Phillips 1969 crop. (1971) 50 early-weaned dairy calves were fed five Stothers and Shebeski (1965) encountered experimental diets at 5 to 7 weeks of age. Ergot palatability problems in feeding triticale of 1962 taken from triticale was added to a complete and 1964 crop years to growing swine. Although barley diet at rates of 0, 0.07, 0.14, and 0.28% the paper states that "ergot has been essentially weight for weight. In the fifth diet triticale contain­ absent from the triticale used in these tests", the ing 0.07 to 0.1 % ergot was used in place of barley. low palatability is now suspected to be at least in Diets were fed ad libitum over 14 weeks; rumen part attributable to ergot in the grain (L. H. samples were taken and ration digestibility was Shebeski 1972, personal communication). determined after 4 and 10 weeks. Diet had no Triticale, ofunstated origin, containing 0.0015°~ significant effect on total volatile fatty acids ergot, was fed to lambs in trials of finishing rations. (VF A), molar percent of VF A, or rumen ammonia Little is known of the effect of ergot on lambs but level. Dry matter and energy digestibilities were the level present did not appear to affect feed lower for the 0.07% ergot diet compared with O/~ intake. Shelled maize was superior to triticale, to ergot, triticale, and 0.28% ergot diets. The effect triticale with maize, and to barley, as judged by on dry matter intake, weight gain and feed weight gain and feed efficiency (Jordan and Hanke efficiency is shown in Table 71. Dry matter intake 1972). of the zero ergot diet was significantly greater than the triticale diet, which was greater than the 0.28% Aflatoxin ergot diet. The zero ergot diet produced signifi­ Toxic substances, known as mycotoxins, are cantly greater weight gains than 0.07% ergot or produced by mould fungi which grow upon many triticale diets, which produced greater gains than grains, grain legumes and oilseeds. The incidence the 0.28% ergot diet. Feed efficiencies were not and control of mycotoxins has been recently significantly different among diets. reviewed by Scott (1973). Of the mycotoxins, the HULSE/LAING: PROTEINS OF TRITICALE 103 aflatoxins, formed by Aspergillus fiarus, are (47%) contained detectable levels, 7 lots (14%) of probably the most important as a hazard to human which were heavily contaminated. health and are certainly those which have been the most extensively studied. Tolerance Levels Kao and Robinson ( 1972) found marked changes in amino acid composition when hard red The reader will note the marked discrepancy spring wheat (var. Selkirk) and hard red winter among different recommended maximum limits wheat (var. Triumph) were inoculated with for ergot infection. The problem of specifying Aspergil/us fiarus spores. The amino acids were limits of tolerance appears with virtually all analyzed by ion exchange chromatography and potentially injurious substances which occur in or aflatoxin by thin-layer chromatography. In the contaminate foods. What constitutes a "safe" sound wheat, the ratio of amino acid to Kjeldahl tolerance limit is influenced by the total quantity protein was about 100%. In mouldy Selkirk the of the food consumed, and the creature for which ratio was 78.86% and in mouldy Triumph, it is intended. Weanlings and young infants are 90.63%. Some amino acids appeared to have been often more sensitive than healthy adults. One can metabolized to form other nitrogenous com­ only urge, in the case of ergot and mycotoxins, pounds still detectable as Kjeldahl nitrogen. In the development of varieties and the implementa­ mouldy Selkirk wheat, cysteine decreased by tion of agronomic, storage, handling, and in­ about 74%, arginine, histidine and glutamic acid spection practices which minimize infection. also decreased. Lysine decreased, but not to such Where there is doubt the advice of competent anextentas in mouldy Triumph wheat. Methionine public or animal health authorities should be was the only amino acid to increase, its increase sought. In any event where tolerance limits are probably being derived from cysteine. In mouldy recommended or prescribed the detailed method Triumph wheat, arginine, histidine and lysine of sampling, inspection and analysis proposed decreased, while methionine increased 35%. The should also be prescribed. amino acid composition of the mould-free wheats were essentially similar. Processing Additives Aflatoxin was detected only in the hard red spring wheat, Selkirk, and practically all was It is recognized that a wide range of additives distributed in the bran. may find their way into processed cereal foods. The incidence ofaflatoxin infection and its level Such additives include flour bleaching and im­ in cereal grains appears to be relatively low, proving agents, dough and alimentary paste compared, for example, with certain oilseeds. conditioners, amylose complexing agents, and Lafont and Lafont ( 1970) examined, chro­ others. Some of these additives affect the biological matographically, contamination with the afla­ value of the protein but no attempt has been made toxin B1 of samples of mixed animal feed from in this publication to review the literature relating two factories in France, and the raw materials of to this subject. which they were composed, taken direct from the In most countries of the developed world there storage silos. are regulations governing the additives which may Detectable levels (above 0.25 µg/kg) ofaflatoxin be included in food intended for human consump­

B1 were found in 15 of the 32 lots of wheat (47%), tion, e.g. Food and Agriculture Organization 7 of the 25 lots of wheat flour (28%), and 4 of the (1959, 1960, 1961a, b, c, 1963a, b, 1969b). The 21 lots of rye (19%). No sample of rye but two lots World Health Organization (1957, 1958, 1961, of wheat (6%) and two lots of wheat flour (8%), 1962, 1964, I 965b, 1966, I 967a, b, 1970, 1972) were heavily contaminated (levels above I 00 has issued monographs on the technology, evalua­ µg/kg). In comparison, 24 of 51 lots of soya cake tion and toxicology of several food additives. •

/ /' ' ...

Tiie prmc1ple o/ tire quem . tire earlil'H /()rm of grain mill. ;,, .Hill fol/011'i'd in many parts a/ th <• dl!vl!loping 11•orld Chapter 5

PROCESSING AND SUPPLEMENTATION WITH OTHER PROTEIN SOURCES

Introduction its distinctive colour. The aleurone layer is important because of its high protein and high Chapter 3 dealt with how the protein content mineral content. and composition of triticale, wheat and rye, are By manual dissection and subsequent analysis, influenced by genetic, varietal, agronomic and Hinton ( 1953) has demonstrated the typically wide environmental factors. The biological value of variation in protein content which occurs among cereal grain proteins can be markedly affected by the various fractions of the cereal grain seeds. what happens to them after they are harvested and Hinton's results, which are based upon a variety in particular by how they are processed. Processing of English wheat, are given in Table 72. From comprehends a wide variety of treatments includ­ Hinton's results it can be seen that the highest ing grinding (milling), mixing with other materials percentage protein levels are to be found in the to form doughs, pastes and slurries, drying, cook­ germ, the scutellum which surrounds the germ, ing and baking. It might be argued that processing and the aleurone layer, and that the protein pro­ should take account of threshing, drying, storage portion declines from the aleurone towards the and chemical treatment of the whole grain. While centre of the endosperm and from the aleurone recognizing the importance of these latter com­ towards the outer seed coats. ponents of the total grain production and uti­ Hinton's general findings are supported by lization system, their impact upon the protein several other investigators. Grewe and LeClerc value is of little consequence except, in a negative ( 1943) analyzed 19 samples of wheat germ from sense, if they are carried out inefficiently and the the USA and demonstrated an average protein grain in consequence suffers significant infection content of 28.9%. Fraser and Holmes ( 1959) and/or infestation. The deleterious effects of analyzed the principal fractions of British wheats infection with ergot and mould-fungi are discussed and found the following average protein contents: in Chapter 4. Insect infestation can result not only endosperm 9.6%, germ 28.5%, bran 14.4%. Kent­ in a gross loss of grain but also in a selective Jones and Amos (1967), from a wide range of nutritional loss in that many insects preferentially samples, quote a spread of wheat germ protein attack the protein-rich moieties of the grain. from 22 to 32%. The basic structure of the wheat grain, which is broadly similar to that of triticale and rye, is illustrated in Fig. 2. The diagram shows that the Milling cereal seed consists at one end of the germ, which The influence of milling on the overall is the embryonic new plant representing 2 to 3% nutritive value of wheat was reviewed by by weight of the total grain, and that the remainder Moran ( 1959) and earlier by Dawbarn ( 1949). consists largely of the endosperm which is the Kasarda et al. (1971) reviewed the composition initial food store for the new plant and forms of the proteins and amino acids of wheat between 80 and 85% of the grain. The germ and fractions, bringing up to date an earlier related endosperm are surrounded by the seed and fruit study by Pence et al. (1964). Lockwood (1960) coats which, when removed, are collectively called reported that the protein content of wheat in the "bran" or "husk". As Fig. 2 illustrates, the typical British bread wheat blends varied between three other layers, collectively called the pericarp, 7.5% and 18%. are the epidermis, the epicarp and the endocarp, It is readily evident that the protein content and they consist largely of cellulose. The episperm and composition of any milled product will be or testa contains the pigments which give the seed influenced both by the protein present in the

105 106 MONOGRAPH IDRC-021e

------Hairs, or beard

BRAN

t------+-- Epidermis f'+---Pericarp Epicarp ~f------Endocarp r1'++++------Episperm or tcsta r.t-++ir-+------Aleurone cells

Ullt-'-:-+------<11'1-Jl+lf-+------Starch cells ~""".,_,"--_____.,._~+4- --- Cellulose cell walls

CREASE

~1~-¥1i;.::..;-..,..,:;.,..,...:...,.;.....:.;""°"~-'-1-++-H------Scutellum :'.':t"~~~~b...:,;_--io...:;....-:r-:~-+--f1-+-t-tt------Plum ule

Dra ...i11g Subi11e Clergn- Vuut:auleurs Fig. 2. Cross-section ofa typical grain of wheat. HULSE; LAING· PROTEINS OF TRITICALE 107

TABLE 72. Protein contributed by wheat fractions (data calculated from Hinton 1953).

Fraction/ Protein/ 100 g whole Protein; Protein/ I00 g protein grain I00 g fraction 100 g grain in whole grain Fraction (g) (g) (g) (g)

Pericarp} 8.0 4.4 0.3 3.5 Testa Aleurone 7.0 19.7 I. 4 15.9 Germ 1.0 33.3 0.3 3.5 Scutellum I. 5 26.7 0.4 4.5 Outer endosperm 12.5 13.7 I. 7 19.3 Middle endosperm 12.5 8.8 I. I 12.5 Inner endosperm 57.5 6.2 3.5 39.9 Whole grain 100.0 8.8 8.8 100.0

original cereal grain and by the proportion of the and shearing stresses encountered in the roller-mill various fractions of the grain which the milled which consequently cause detachment and partial product contains. disintegration of the endosperm particles initially The milling of cereals into flour is probably attached to the bran. man's most ancient food processing industry. The purpose of most roller-milling operations is Primitive man pounded the cereal seeds between to separate the endosperm from the bran and germ rocks until, about 4000 B.C. the more efficient and to convert it to a fine white flour or, in the case hand quern appeared. Within the ancient quern of durum wheats intended for alimentary pastes, are to be seen the first traces of modern milling into clean, uniform, yellow particles called semo­ techniques in that rotary stones, carved with lina. The percent proportion by weight of flour radial grooves, revolve one upon the other thus obtained from a given quantity of wheat or other breaking open the grain and freeing the white cereal grain is known as the extraction rate. Thus starchy endosperm which is then ground and a 50% extraction rate would contain 50% by sieved to produce the fine white powder known as weight of the grain milled and would consist of "flour" which was from earliest times described as almost pure endosperm. A 100% extraction rate the "flower" (the finest portion) of the grain. would consist of the whole ground grain.

ROLLER MILLING EXTRACTION RATE Lockwood ( 1960) and Ziegler and Greer ( 1971) Ziegler and Greer ( 1971) present data which have presented comprehensive reviews of modern illustrates the relation between extraction rate roller-milling technologies. Briefly, a roller-mill and protein content. Table 73, derived from Ziegler consists of two series of paired rolls, (a) the and Greer, indicates the contrast in protein content break rolls which are corrugated and counter­ between low (76% and below) and high (above rotate at different angular velocities thereby 80%) extraction rates. shearing and breaking the cereal grains and In producing low-extraction flours, the protein­ permitting the endosperm to spill out in large rich aleurone layer is virtually eliminated with the chunks, (b) the reduction rolls, which are smooth, bran. The 85% extraction flour would contain or very finely corrugated, and crush or reduce the most of the aleurone layer and therefore would endosperm to fine particles. Interspersed with the retain between 96 and I 00% of the protein flow of broken and crushed wheat particles between originally present. The question of what constitutes the break and reduction rolls one finds a series of an optimum percentage extraction rate needs to sieves, gravity tables and aspirators which separate be carefully considered, particularly in the less particles of different size, density and effective developed countries, several of which convert mass. The comparative toughness of the bran imported wheat to low-extraction rate flours for enables it to withstand the application of crushing local consumption and export the separated bran 108 MONOGRAPH IDRC-02le

TABLE 73. Compositions• of flours of various extraction rates in series milled from given wheats (data from Ziegler and Greer ( 197 l ).

Extraction rate of flour (%)

42--46 65 70 75 80 85 100

l l. 9 12.9 13.2 13.4 13.7 13.8 Protein (N x 5. 7), % 7.73 8.01 8.08 8.31 8.65 9.0 (at 14% moisture) 10.7 10.9 l l. 6 12.0 12.0 n 11. 3 l l. 9 12.4 12.3 86.6 93.6 95. 7 97. l 99.5 100.0 Protein (N x 5.7), as 85.9 89.0 89.8 92.4 96. l 100.0 % of protein in wheat 89.2 90.8 96.6 100.0 100.0 n 92.0 96.8 100.0 100.0 "Flour A: Manitoba wheat: laboratory milled Flour B: English wheat: laboratory milled Flour C: Manitoba 65%, English 35%: laboratory milled Flour D: Mixed wheats: commercially milled. and germ to developed countries for use m containing 18 to 20% protein can be obtained by animal feeds. fine grinding and a single air classification. In the

AIR CLASSIFICATION authors' experience the most extreme result was The endosperm of wheat consists of starch obtained from a sample of Gaines wheat in which granules of various sizes embedded in a matrix of the original 75% extraction flour contained 8.5% protein. There is a limit to the degree of fineness protein. After pin-mill grinding and fractionation in wheat flour which can be produced by break in an Alpine air classifier, the fraction below an and reduction milling. If the reduction rolls are average diameter of 17 microns was found to con­ set too close together the particles are simply tain an average of 24% protein (Hulse, 1966 compressed into flat discs. By employing impact­ unpublished data). milling, as exemplified by the pin disc-mill, ABRASIVE DECORTICATION the endosperm particles can be reduced to flours An alternative principle of milling, which of extremely fine particle size. During such fine deserves more attention than it has received, con­ grinding much of the endosperm protein is sists of abrasive decortication in which the seed liberated as free protein in the form of small coats (bran) are removed by passing the grain wedge shaped slivers. Since these fine protein between moving abrasive surfaces. Such a process fragments possess the smallest average effective has been described by deMan et al. (I 973). mass, they can be separated from the starch-rich The theoretical advantages of such a system are fraction by air classification in which a centrifugal that the proportion and thickness of bran coats force is counteracted by a centripetal air drag. removed can be varied according to the intensity The relative magnitude of these opposing forces and duration of the abrasive forces applied. Ideally determines the critical particle size or effective one could remove the high cellulose epicarp and mass at which the flour will be fractionated (Jones testa without removing the high protein aleurone and Halton 1958; Jones et al. 1959). The finest layer thus leaving perhaps more than 95% of the particles of lowest effective mass will be richest in original grain seed protein intact. Furthermore, protein. certain of the antimetabolites and microorga­ Because of the physical nature of their endo­ nisms present in and on the outer seed coats sperm, soft wheats respond in greater degree to would also be removed. Preliminary evidence from fine grinding and air classification than hard an IDRC project in West Africa suggests that wheats. Starting with Ontario soft wheat flours grain processed by abrasive decortication as of roughly 9% average protein content, fractions described by deMan et al. (I 973), and subsequently HULSE/LAING: PROTEINS OF TRITICALE 109 converted to flour, is more stable in storage, The four basic and essential ingredients of probably because most of the mould-fungi present leavened bread are flour, salt, yeast and water. on the seed coats are removed with the bran. Leavened bread comes in a remarkable range of During break milling or roller milling the micro­ compositions, weights, shapes and sizes. organisms on the outer seed coats tend to be In the Near East, thin discs of fermented dough homogenized with the ground whole grain are baked for a few minutes at temperatures close particles. It would be worthy of experiment to to 400°C which causes the dough to rise rapidly discover if both ergot, where present, and some almost to the conformation of a hollow sphere of the antimetabolites present in the outer seed (Dalby 1969). coats described in Chapter 4, might be removed by On the Indian sub-continent chapatis are made abrasive decortication with a consequent improve­ from unfermented, unleavened discs of dough ment in the quality of the grain. baked upon a hot plate (Chowdhary 1954, Dharam Jit Singh 1956). RYE MILLING The influence of milling upon the protein value Supplementation with Other Protein Sources of rye grains appears to have received scant atten­ In making bread, wheat or other cereal flour can tion though some of the information is to be found be supplemented with a very wide range of protein in Kent (1966) and Kent-Jones and Amos (1967). sources and other nutrients. Hulse (in the press) In Canada rye is normally milled by a break and has reviewed the technological and nutritional reduction process to produce two main products, implications of supplementing bread and other dark rye flour which consists essentially of the baked cereals with various protein sources. whole rye grain, and light rye flour of approxi­ At the end of this Chapter, two aspects of mately 70% extraction. One set of commercial protein supplementation are considered: (a) that data indicates that the protein content of light which improves the amino acid balance and (b) rye flour is roughly 25% lower than that of dark that which increases the total proportion of protein rye flour milled from the same grain. nitrogen. Since, in much of the work reported, the two aspects are virtually inseparable, the section Secondary Processing has been organized according to the sources of protein supplementation rather than according The secondary processing of cereal grains con­ sists in the main of cooking or baking the milled to the primary purpose of supplementation. The products with water and other ingredients. The somewhat lengthy review of supplementation is, simplest processing consists of cooking in steam the authors believe, relevant particularly to the or water to produce a gruel, porridge or granular needs of developing countries. In several of these countries, where the demand for bread is steadily material of which couscous is typical (Ferhi 1970). increasing (the annual increase exceeds 8% in In the making of alimentary pastes such as Africa) one encounters a growing interest in macaroni, noodles and spaghetti, the cereal flour bread made from composite flours in which wheat is combined with water and sometimes other flour is supplemented with other cereal flours, ingredients, before being dried or sheeted and cut into thin strips or shapes and dried. Alimentary including maize, sorghum and millet, with root paste production is discussed by Hummel (1966), starches, such as cassava, plus legume proteins Irvine (1971) and Winston (1971). (Food and Agriculture Organization 1970c; Hulse (in press)). This is a subject that will doubtless BAKING receive increased attention as more countries seek Baking is perhaps the oldest and most widely comparative self-sufficiency in cereal grains. The practised method of processing cereal grain flours. performance and acceptability of composite flours The principles and practices of baking bread, cakes containing triticale is being studied at the Uni­ and biscuits are the subject of many publications, versity of Manitoba. e.g. Pyler (1952), Calve) (1962), Fance (1968), This Chapter is concerned primarily with how Fance and Wragg (1968), Kiger and Kiger (1968), the biological value of the protein of triticale, Matz (1968, 1972), Sultan (1969), Pomeranz and wheat and rye may be influenced by primary and Shellenberger (1971 ), Smith (1972), Bennion and secondary processing and by supplementation. Bamford (1973). Not surprisingly in view of its long history as a 110 MONOGRAPH IDRC-021e

TABLE 74. Generic identification key of wheat, rye and triticale varieties (Unrau and Jenkins 1964).

Ploidy Designation Botanical Description

Tetraploid 48113 Triticum turgidum var. ramoso mega/opo/itanum Tetraploid 48121 T. turgidum var. p/inianum Tetraploid 48249 T. po/onicum var. /erissimum Tetraploid 48276 T. persicum var. stramineum Tetraploid Ramsey (durum wheat) T. durum var. hordeiforme Tetraploid Stewart (durum wheat) T. durum var. hordeiforme Tetraploid Fourex (rye) Seca/e cerea/e (induced tetraploid) Hexaploid Pembina (bread wheat) T. aestirum var. /utescens Hexaploid Selkirk (bread wheat) T. aestirum var. /utescens Hexaploid 6A49 (bread wheat) T. aestirum (synthesized from T. turgidum var. nigro-baratum x Ae. squarrosa) Hexaploid 6A20. l (Triticale) T. durum "Carleton" -S. cerea/e Hexaploid 6A66.l (Triticale) T. dicoccoides-S. cerea/e Hexaploid 6A67. l (Triticale) T. persicum-S. cerea/e Octaploid 8Al 12 (Triticale) T. aestirum-S. cerea/e (awnless) Octaploid 8Al25 (Triticale) T. aestil·um-S. cerea/e

staple of man's diet, a formidable body of litera­ The high crude protein content of the triticale ture can be found which relates to bread and other grain was considered to be associated to some baked products from wheat flour and to a much extent with the rye genomes, as was, also, the lesser extent to bread made from rye flour. At the shrivelled kernel characteristic. time of writing, comparatively little has been Vaisey and Unrau (1964) in a paper published published concerning bread and other cereal foods before Unrau and Jenkins (1964) but describing from processed triticale. Nevertheless, it is believed later work, examined the same varieties primarily that much of what is known concerning the in­ for their sugar content. The values obtained for fluence of processing and supplementation upon protein were in close agreement with those the nutritional value of wheat and rye products described by Unrau and Jenkins (1964). may be extrapolated to triticale. It is for this Berry et al. ( 1971) compared a hard red spring reason that the following rather extensive chapter (HRS) flour, durum semolina and hexaploid is included in this publication. triticale, all grown in North Dakota and milled on a Buhler mill, and a commercial rye flour. They were primarily concerned with the characteriza­ Milling tion of the triticale starch but the protein contents Triticale were determined using the official methods of the Unrau and Jenkins (1964) investigated several American Association of Cereal Chemists ( 1962). triticale varieties, both hexaploid and octaploid, Expressed on a dry matter basis, the percent together with species of wheat and rye. A generic protein contents were: HRS flour 16.1 ; durum description of the varieties used is given in semolina 17 .5; rye flour 11.0; triticale 12.4. Table 74. Lorenz and Maga ( 1972) investigated the com­ Grain samples were milled in a Buhler laboratory positions of fatty acids, carbonyls and hydro­ mill. The hexaploid triticales were passed through carbons in eight samples of grain all grown on the mill stream a second time to obtain a higher irrigated sites at Fort Collins and Center, Colorado extraction. Table 75 records the 1000-kernel in 1971. Two winter wheats (Scout and Caprock), weight, the crude protein of the seed, on a dry two spring wheats (lnia Res and Ciano Sib.), two weight basis, the flour yield, and the flour protein spring hexaploid triticales (6TA204 and 6TA206) also o~ a dry weight basis. Protein content was and two winter hexaploid triticales (TR385 and determined by Kjeldahl procedure (Association TR386) were tempered and milled on a Brabender of Official Agricultural Chemists, 1950). Quadrumat Junior mill. Moisture and ash were HULSE/LAING: PROTEINS OF TRITICALE Ill determined by the methods of the Association of TABLE 76. Characteristics of wheat and triticale culti­ Official Agricultural Chemists (1960); protein vars (14% moisture basis) (Lorenz and Maga 1972). content by the Udy Protein Analyzer method (American Association of Cereal Chemists 1962). Grain Flour The grain protein, flour protein and flour ash of protein protein Flour the cultivars examined are given in Table 76. (N x 6.25) (N x 5.7) ash Madi and Tsen ( 1973) determined the pro­ (%) (%) (%) teolytic activity by a modified Ayre-Anderson Spring triticale method (McDonald and Chen 1964; Wang and 6TA204 14.6 11. 3 0. 53 6TA206 14.5 11. 7 0.54 TABLE 75. 1000-kernel weight in grams, crude protein Winter triticale of seed. flour yield, and flour protein of varieties TR385 12.0 8.7 0.47 examined (data from Unrau and Jenkins 1964). TR386 10.8 8.6 0.47

1000- Crude Spring wheat kernel protein Flour Flour Inia Res. 12.7 12.4 0.46 weight of seed yield protein Ciano Sib. 13.2 12.5 0.48 Variety ~) (%) (%) (%) Winter wheat Scout 12.0 11.6 0.43 Durum wheat Caprock 11.2 11.0 0.41 Ramsey 43.7 16.5 74.1 15.7 Stewart 51.8 16.0 72.3 14.4

Tetraploid wheat Grant 1969; Bushuk et al. 1971) of six triticales, 48113 44.3 15. 0 63. 3 14.2 three ryes and four wheats milled on Brabender 48121 44.6 15 .0 67. I 14.1 Quadrumat and Miag experimental mills. The 48249 67.2 18. 3 71. 8 I 17. grain protein (N x 5.7, dry basis), flour yield, 48276 29.3 17. 9 63.1 15.5 flour protein, flour proteolytic activity, bran Bread wheat protein and bran proteolytic activity of the samples Pembina 29.7 17.2 69.6 16.2 are shown in Table 77. The Mexican triticales, Selkirk 36.9 17. I 71. 9 16.3 Armadillo and Bronco, showed higher protein 6A49 44.6 18.0 65.6 17.2 contents and flour yields than the Kansas grown Rye varieties. The proteolytic activity of triticale was Fourex 36.9 19. I 59. 7 13.6 comparable to that of rye which was significantly higher than that of wheat. The proteolytic activities Hexaploid triticale of the brans were considerably higher than those 6A20. I 46.3 19. 8 63. 7 18.8 6A20.2 44.0 19.5 63.0 17.3 of the flours. No direct correlation between 6A66. I 45.6 19.3 62.7 17. I protein content and proteolytic activity was 6A67.1 43.6 17.3 60.4 15. I observed. Moisture, protein contents and pro­ 6A67 .2 42.5 17. I 63 .4 15.3 teolytic activity of the milling fractions of a Kansas 6A67 .3 41.8 17. 3 61. 5 16.2 triticale are shown in Table 78. The proteolytic 6A67 .4 44.1 17.2 64.9 15.3 activity of the fractions increased as the protein 6A67 .5 43.7 16.9 66.7 15.3 content of the fractions increased. 6A67 .6 42.2 17.3 66.8 15.5 Chen and Bushuk (l 970a, b, c) reported on the 6A67. 7 43. I 16.6 66. I 15.2 solubility characteristics and amino acid com­ 6A67 .8 42.4 16. 8 66. 8 15.2 position of one line of triticale and of its durum 6A67 .9 41. 8 17. I 68. 8 15.5 6A67. IO 42.0 17.3 66.6 15. I wheat and rye parents. Included in the trials was a 6A67. I I 43.4 16. 3 63 .4 15. I cultivar of hard red spring wheat, Manitou. The 6A67 .12 44.5 18.1 64.5 16.9 triticale, 6A 190, was originally produced at the University of Manitoba by crossing the durum Octaploid triticale wheat variety Stewart 63 and the rye variety 8AI 12 26.1 17.9 64.7 16.6 Prolific. With the hard red spring wheat, Manitou, 8Al25 38.8 19.2 67.5 18.0 the four species were grown together in 1966 at 112 MONOGRAPH IDRC-021e

TABLE 77. Relative proteolytic activities of flours and brans of wheat, rye, and triticale (data from Madi and Tsen 1973).

Whole Grain Flour Bran

Protein Protein Activity Protein Activity (N x 5.7 Flour (N x 5.7 (mg protein (N x 5.7 (mg protein dry basis) yield dry basis) per ml dry basis) per ml (%) (%) (%) supernatant) (%) supernatant)

Wheats Eagle (Manhattan, 1972) 12.7 70 11. 8 I. 74 14.3 4.31 Centurk (Manhattan, 1972) 12.7 73 11.4 I. 75 15.2 4.87 Scout-I (Garden City, 1971) 13.6 72 13. I I. 73 16.7 5.37 Scout-M (Manhattan, 1971) 14.6 72 13.9 I .65 17.7 4.80 Ryes Minn. 11 (Columbia, Mo. 1972) 10.0 51 7. I 2.61 13.9 6.48 Balbo (Manhattan, 1972) 12.6 45 8. I 2.07 15.7 5.77 Balbo (Manhattan, 1971) 16.3 48 12.7 2.08 19.5 5.75 Triticales Triticale-1 (Manhattan, 1971) 13.7 64 12.6 I. 98 18. I 6.60 Triticale-F (Garden City, 1971) 15.1 61 14.1 2.01 18.6 6.15 Triticale-385 (Garden City, 1971) 15.4 62 13.6 2.00 20.0 5.70 Triticale-298 (Manhattan, 1971) 17.5 58 15.5 1.93 21.0 5.96 Armadillo (Mexico, 1971) 16.2 67 13.9 2.68 21. 3 6.35 Bronco (Mexico, 1971) 19.9 69 17.6 2.53 23.3 6.45 the University of Manitoba. The samples were acids from the rye sample was rather low, for milled into flour on a Buhler experimental mill reasons which were not investigated. after tempering overnight to 16.5% moisture. The Anderson et al. ( 1972) reported on the effect of flours were characterized using the standard milling and air classifying a triticale (FasGro 204). methods of the American Association of Cereal The composition of the triticale grain on a mois­ Chemists (1962), unless otherwise specified. Pro­ ture free basis is given in Table 81. tein contents (N x 5.7) were obtained using a Using a Buhler pneumatic laboratory mill a semi-micro Kjeldahl method and amino acids by straight grade flour was obtained from this triti­ autoanalyzer The 1000-kernel weight and mois­ cale of about 64% extraction, flour protein 12.1 % ture content of the grain of the four varieties with and ash 0.54%. both on a 14% moisture basis. their flour yield, ash, moisture and protein content Bran and shorts were separated from the flour are given in Table 79. The amino acid compositions without difficulty. A second straight grade flour, of the flours from the four varieties are given in 12.7% protein and 0.57% ash was ground more Table 80. finely by three passes through an Alpine Kolloplex In general Chen and Bushuk (l 970a) found the mill (Model 16 oz). The finely ground flour was air amino acid composition of the triticale to be inter­ classified in a Pillsbury laboratory model classifier. mediate between its parent species, durum wheat Yields of the air classified fractions and their and rye. They comment that the recovery of amino analyses are given in Table 82. HULSE;LAING: PROTEINS OF TRITICALE 113

TABLE 78. Proteolytic activity in various miIIing Anderson et al. (1972) also wet-milled triticale fractions of a Kansas triticale (data from Madi and grain by a laboratory procedure (Anderson, R. A., Tsen I 973). 1963). No particular problems were encountered, the triticale responding in a manner similar to soft Proteolytic wheats. Starch and gluten were separated by the Protein activity Martin, or dough ball process (Knight 1965) but (N x 5.7 (mg protein the softer nature of the gluten presented difficulties Moisture dry hasis) per ml in the washing process. MiII fraction (%) (~o) supernatant) Hexaploid triticale, var. Cachirulo, provided by Straight grade 12.7 11. 2 I. 78 the (Spanish) lnstituto Nacion al de Investigaciones I st break 14.5 7.5 I. 33 Agronomicas, was test milled using a Buhler model 2nd break 14.0 9.1 I .43 M.C.K. and a Miag model Vario C ex 2. The 3rd break 12.6 12.6 2.07 authors (de la Plaza et al. 1969) suggest a milling I st middlings 13.0 12.0 I. 55 diagram from which six final products may be 3rd middlings 11.6 15.5 2.80 obtained, two flours, two protein concentrates 4th middlings 10.3 16.5 3.31 and two brans. Four of these fractions, two flours 5th middlings 9.5 18.0 4.16 and two protein concentrates obtained from a Reduction shorts 8.2 14.8 6.10 combination of the Buhler and Miag Vario experi­ Red dog 9.0 15.6 6.40 Break shorts 11.0 18.4 6.43 mental mills were compared with a triticale flour Germ 9.6 21.0 5. 20 obtained from a normal milling on the Buhler and Whole grain 9.7 13.3 a similarly Buhler milled control wheat flour obtained from a blend of I 0% Florencia Aurora, I 0% Impeto, 30% Aragon 03 and 50% Negrillo Anderson et al. (1972) also quote fractionation (Vallejo et al. 1969). The chemical characteristics results for triticale compared with those found are given in Table 84. Nitrogen was determined by by other authors for rye and wheat (see Table 83). Kjeldahl, ash by the method of the American It is interesting to note that whereas the yields Association of Cereal Chemists ( 1962). The of recovered fractions from triticale more closely methods for fat and fibre determination were not resemble those from rye than from wheat, the total identified. protein shift is significantly higher in the case of triticale than the other two cereals. Total protein Wheat Flour shift is defined by Anderson et al. (1972) as the Table 3 in Chapter I shows average differences sum of the protein shifted into the high-protein and ranges in amino acid composition among fractions and out of the low-protein fractions, whole wheat, germ, bran and flour of 70 to 80% expressed as percentage of the total protein present extraction rate. The data in this table cannot be in the flour, as described by Gracza (1959). directly related to each other since they are

TABLE 79. Grain, I 000-kernel weight and moisture content; flour yield, ash, moisture and protein of triticale, its parental species and a hard red spring wheat (Chen and Bushuk I 970a).

Durum Hard red Triticale Spring rye wheat spring wheat 6Al90 Prolific Stewart 63 Manitou

Grain JOO-kernel weight, g 43.4 28.6 44.9 30.6 Moisture content, % 14.0 13.0 13.7 13.5 Flour Yield,% (14% moisture basis) 58.9 59.6 67.7 71. 7 Ash content,% (14% moisture basis) 0.42 0.68 0.66 0.48 Moisture content, % 14.0 13.0 13.7 14.8 Protein content, % ( 14% moisture basis) 9.8 9.9 12. I 13.6 114 MONOGRAPH IDRC-02le

TABLE 80. Amino acid compositions of flours from rye, triticale, durum wheat and hard red spring wheat (micromoles amino acid per milligram nitrogen in sample) (Chen and Bushuk 1970a).

Rye Durum HRS wheat Amino acid (Prolific) Triticale (Stewart) (Manitou)

Aspartic acid 2.41 2.23 1.89 1.47 Threonine 1.29 1.34 1.29 1.20 Serine 1.84 2.17 2.26 2.21 Giutamic acid 9.77 11. 7 13.2 15.7 Pro line 6.26 6.44 6. 56 6.21 Glycine 2.40 2.70 2.54 2.73 Alanine 2.15 2.08 1.99 1.84 Valine 2.10 2.20 2.21 2.11 Cystine 0. 52 0.70 0.60 0.59 Methionine• 0.68 0.79 0.78 0.65 lsoleucine I. 51 I. 74 I. 87 I. 71 Tryosine 0. 56 0.81 0.80 0.78 Phenylalanine 1.47 I. 58 1.80 2.10 Ammonia 9.41 10.0 12.5 13.6 Lysine 1.12 1.00 0.85 0.82 Histidine 0.65 0.79 0.83 0.85 Arginine I. 23 1.43 1.13 1.20 Tryptophan not analyzed N recovery, % 76.2 84.5 87.8 91. 7

•Including methionine oxide.

TABLE 81. Composition of triticale (FasGro 204) derived from several sources, but it can be ob­ (moisture free basis) (data from Anderson et al. 1972). served from the table that, compared to wheat, the contents of lysine, arginine, alanine, aspartic Crude protein (N x 5. 7), % 17. I (ca 14.6 at 14% H20) acid and glycine are lower in flour and higher in Crude fat,% I. 7 bran and germ, while the contents of glutamic acid Crude fibre, % 3.1 and proline are higher in flour and lower in bran 2.1 Ash,% and germ. This is a reflection of the relatively high Nitrogen-free extract, % 76.0 amounts of endosperm protein present in com-

TABLE 82. Fine grinding and air classification of triticale flour (14/;, moisture basis) (data from Anderson et al. 1972).

Kind of flour Mass median and Yield Protein Ash Fat Maltose diameter fraction number (/;,) (/;,) (/;,) (%) values µ

Straight flour 12.7 0. 57 0.69 161 29 Reground flour (3 passes) 12.0 0.59 0.62 246 20 Fraction I 10. 7 34.3 I. 15 I. 56 372 11 Fraction 2 8. 0 28.4 0.88 1.13 374 12 Fraction 3 6. 4 20.7 0.73 0.85 359 15 Fraction 4 22. 8 6.7 0.50 0.41 187 18 Fraction 5 14.0 4.8 0.45 0.34 184 21 Fraction 6 16. 8 4.4 0.42 0.30 157 23 Fraction 7 11. 7 5.6 0.46 0.31 149 25 Fraction 8 (coarse residue) 9. 6 13.3 0.50 0.42 204 31 100.0 HULSE/LAING PROTEINS OF TRITICALE 115

TABLE 83. Comparison of fractionation responses of reground flours from triticale, rye, and wheat ( 14% moisture hasis) (Anderson et al. 1972).

Rye• Wheat

Hard Hard Commercial Durum Red Spring Red Winter Sample Triticale Balboa Mix Wellsb Selkirkb Wichita'

Straight flour, protein,% 12.7 8.2 10.9 15.7 12.8 10.9 Maximum range of protein contents, % 34.3-4.4 18.6--3.4 21. 5-6. 2 26. 2-11 .4 23.7-7.6 29.4-5.5 Combined high-protein fractions 1-3 Protein,% 29.0 15.9 19.8 23 .6d 19.4 24.4 Yield,% 25. 1 26.8 25.8 9. 2d 18.8 21.0 Combined starchy fractions 4-7 Protein,% 5.5 4.8 7.4 12.0' 8.7 6.5 Yield,% 65.3 60.4 56.8 30. 3' 49.5 52.9 Coarse residue fraction 8 Protein,% 13.3 5.2 9.6 16.4 14.2 9.8 Yield,% 9.6 12.8 17.4 42.2 31. 7 26.1 Protein shifted, total, % 73.0 50.0 41.0 15.0 29.0 50.0

•Unpublished data. bPeplinski et al. ( 1965). 'Stringfellow and Peplinski ( 1966). dCombined fractions 1 and 2. 'Combined fractions 4, 5, and 6.

TABLE 84. Characteristics of triticale milling products and control wheat flour (dry maller basis) (data from Vallejo et al. 1969).

Triticale Wheat blend Buhler milled Flour Flour Concentrate Concentrate Buhler milled flour 1 2 2 flour

Ash,% 0.67 0. 59 0.77 1. 38 1. 78 0. 58 Protein (N x 6.25), % 18.2 16.2 19.3 22.2 24. l 10.6 Fat,% 1. 5 1.0 1.0 2. 1 3.0 1. 2 Fibre,% 0.6 0.7 0.8 0.9 1. 8 0.4 mercially milled flour, from which bran and germ (1957, I 960b); Horn et al. (1958); Bruggemann are customarily excluded. and Erbersdobler (1967); Kohler and Palter Kasarda et al. ( 1971) in their review of the amino (196 7); Waggle et al. (196 7); Tkach uk and Irvine acid composition of wheat flour provide tables (I 969a). A similar pattern emerges to that shown which compare the amino acid composition of in Table 3. However, soft wheats, comparatively flour fractions with the wheats from which they are low in protein, tend to have a higher content of milled citing, amongst others, Hepburn et al. lysine, arginine and other essential amino acids 116 MONOGRAPH IDRC-02le

TABLE 85. Relative nutritive value (RNV) of wheat 65%. Tryptophan was not determined. The con­ and wheat products (as percentage of" RNV ol lactal­ tent of the essential amino acids was not greatly bumin ±the standard error) calculated using rat weight affected by the milling process, except for lysine, gain and body water as measures of response (data which was materially decreased by milling to 75% from Miladi et al. 1972). extraction and showed a further decrease on milling to 65% extraction rate. Based on Based on Miladi et al. (1972) examined germ, wheat weight gain body water protein concentrate, red dog, bran, shorts, Wheat germ 80±5 79±3 patent flour and clear flour obtained by standard Wheat protein concentrate 72±4 75±3 milling procedures from a single sample of hard Red Dog 60±3 60±3 red spring wheat. The amino acid content was Shorts 55±3 57±2 determined by autoanalyzer. Protein availability Bran 49±3 51 ±2 was determined by an in vitro method (Kohler Whole wheat 36±3 39±2 et al. 1969). The relative nutritive value (RNV) Patent flour 23±4 26±3 was determined as described by Hegsted and Chang Clear flour 25±3 27±2 (l 965a, b), Hegsted and Worcester (1966), The mean value ± 2 standard errors gives the 95~0 Hegsted et al. (1968), using young male rats confidence limits of each value. weighing approximately 50 g. The RNV is defined as the slope of the dose-response curve of the than do strong (higher protein) wheats (Kasarda protein under test divided by the slope of the dose­ et al. 1971 citing Lawrence et al. 1958; Hepburn response curve obtained with the standard protein, and Bradley 1965; and Waggle et al. 1967). lactalbumin. Body-weight gain in the 3-week Soft wheats are customarily used more exten­ experimental period and the body-water of the sively in cakes and biscuits (cookies) than in animals at the end of the experiment were used as bread flours. However, recent technological de­ measures of "response". velopments, including the Chorleywood Bread The RNV of the various fractions is given in Process (Chamberlain et al. 1966) have made it Table 85. Patent flour has the lowest, and germ possible to include significantly larger proportions the highest, RNV, and the RNV correlates highly of soft wheat flours in bread than heretofore. with the lysine content of the proteins of the Nunnikhoven and Bigwood (1959) reported on fractions analyzed. The essential amino acid the amino acid composition (ion exchange chro­ contents are shown in Table 86. A chemical score matography) of a sample of Manitoba No. 2 based on the essential amino acid content of egg wheat, and of the flour commercially milled from protein did not predict the RNV. The total it in Belgium at two extraction rates, 75% and protein content and the digestibility of the fractions

TABLE 86. Essential amino acid composition of wheat mill fractions (in grams amino acid/16 g N) (data from Miladi et al. 1972).

Clear Patent Whole Red Wheat Flour Flour Wheat Dog Bran Shorts wpc• Germ

Lysine 1.88 I. 95 2.47 3.78 4.10 4.21 4.59 4.92 Tryptophan I. 11 1.10 I. 28 I. 38 I. 92 I. 53 I. 35 I. 28 Threonine 2.62 2.67 2.82 3.24 3.29 3.31 3.49 3.52 Valine 4.53 4.58 4.88 5.16 5.17 5.18 5.40 5.22 Jsoleucine 4.08 4.30 4.01 3.83 3.63 3.61 3.83 3.68 Leucine 6.78 6.78 6.54 6.30 5.98 5. 96 6.26 6.04 Tyrosine 2.93 3.11 2.89 2.69 2.66 2.66 2.78 2.58 Phenylalanine 4.99 5.08 4.73 4.29 3.91 3.81 4.02 3.78 Cystine 2.76 2.64 2.44 2.36 2.29 2.24 2.21 1.88 Methionine 1.88 I. 97 1.89 I. 92 I. 74 I. 81 2.13 I. 89

•wheat protein concentrate. HULSE/LAING. PROTEINS OF TRITICALE 117 as measured by the in vitro method of Kohler TABLE 87. Total protein content and digestibility of et al. (1969) are given in Table 87. the protein in wheat fractions. in vitro method of The effect of varying extraction rates of wheat Kohler et al. 1960 (data from Miladi et al. 1972). flour on the nutritional value of the breads made from the flours was described in the classic Protein experiments of Widdowson and McCance ( 1954) content Digesti- (N x 6.25) bility in feeding undernourished European children. Wheat fraction (%) (%) Contrary to what had been expected from rat feeding trials, no significant differences were found Wheat germ 25.25 91.6 between the breads, which supplied about 75% of Wheat protein concentrate 27.40 96.5 the calories in diets which were otherwise poor. Red dog 17.40 94.1 Hughes ( 1955) estimated the amino acid content of Shorts 19.70 83.9 the different diets used and from this it appeared Bran 17.20 69.4 that the lysine content in the diets was adequate, Whole wheat 16.67 91 .0 and that this amino acid was therefore no longer Patent flour 14.39 99+ limiting. Under these conditions the differences in Clear flour 19 .15 99+ extraction rate would not be of nutritional significance. percentage of the nitrogen absorbed was approxi­ Hutchinson et al. ( 1956) fed male weanling rats mately 29% for the wholemeal, 25% for the 85% with bread made from wholemeal flour and from extraction flour and 21 % for the 50% extraction 70% extraction flour, laboratory milled from a flour. clean sample of Manitoba No. 2 wheat. When, in The whole wheatmeal, and seven of the 42 the diets, the lysine content of both the wholemeal milling fractions obtained from it at a com­ and white bread was raised to about I%. lysine mercial mill in Sweden, were fed to provide 9.4% was no longer limiting and the rats grew equally protein in the diet of rats and mice in trials reported well on the lysine supplemented diets. Hutchinson by Munck ( 1972). The fractions fed included the et al. found no evidence of any other amino acid embryo, P42, two fractions of the inner endosperm, becoming limiting, once the lysine requirement had PIO and Pl2, and four fractions from the outer been satisfied. Weanling rats appear to be endosperm, P37, P38, P39 and P41. The composi­ extremely sensitive to small changes in the lysine tion of the fractions in protein, lysine, fibre, ash content in diets of which the lysine content is and resorcinols is given in Table 88. within the range of0.3 to 0.5%. The nitrogen retention (NR) in mice and the Gontzea et al. (1970) reported on rat feeding net protein utilization (NPU) in rats were reason­ trials in which wheat wholemeal and flours of ably well correlated. There was a marked negative 85%, 75%, 50% and I 0% extraction rates were relationship between crude fibre content and true compared. Male weanling Wistar rats, in groups digestibility for rats. Because of the high degree of IO, were pair-fed the flours, as such, with only of correlation between crude fibre and digesti­ the addition of 2% liver oil, over 30 days. Growth bility, and between resorcinol content and digesti­ rate, food consumption and protein efficiency ratio bility, it was not possible to differentiate statistically were measured. Growth rate was highest on the between the effects of these two factors. wholemeal diet and fell as the extraction rate was The high lysine P37 fraction gave an NPU in reduced; the most important reduction, statisti­ rats equal to that of the P42 germ diet; in mice cally, was between flours of 85% and 75% extrac­ the P37 diet gave a higher NR than did the P42 tion rates. Flours of 75% and 50% extraction rate diet. In the mouse trials, the P3 7 to 41 diets were twice, and those of 10% extraction rate, five showed a continuously decreasing gain in protein to eight times, worse than the 85% extraction rate with decreasing consumption of lysine. The wheat and wholemeal as measured by growth rate. germ diet lies on the upper side, and the low­ In nitrogen balance tests with adult rats, pair­ lysine PIO, Pl2 and the whole wheat (W) diets on fed, the digestibility coefficient of the 50% extrac­ the lower side, of the regression line of the tion flour exceeded 90%, of the 85% extraction P37 to 41 diets. The digestible lysine in the germ flour was approximately 85%, and of the whole­ was thus utilized less effectively, and the PIO, Pl2 meal 78%. However, the nitrogen retained as a and W diets more effectively, than expected when 118 MONOGRAPH IDRC-02le

TABLE 88. Protein, lysine, fibre, ash and resorcinol contents of commercial milling fractions of a Swedish wheat (data from Munck 1972).

Crude protein Resorcinol (N x 6.25) Lysine Fibre Ash relative (%) (g/16gN) (%) (%) units

PIO} inner 10.0 2.31 0.10 0.24 14 P 12 fractions 14.2 3 .11 0.90 1.80 39 19.3 6.01 4.70 3.52 124 P37}P 38 outer 14.8 5.88 9.30 3.93 382 P 39 fractions 14.0 5.66 11.60 5.10 516 P41 12.8 4.86 12.75 6.44 538 P 42 embryo 30. l 7.70 2.40 4. 55 40 Whole meal 10.7 3.34 I. 95 1.28 90 compared with the results of the P37 to 41 Tara et al. ( 1971) examined 33 samples of flour fractions. from Indian commercial roller mills. A composite The addition of bran to a diet based on white grist was prepared by mixing equal amounts of flour improved the growth rate and food con­ wheat, mainly US red wheat, from the mills, sumption of weanling rats partly, it is suggested, conditioning the mix to 15 .5% moisture and milling because of the supplementary effect of the higher to give a 69.5% extraction flour. Lysine, threonine lysine in the bran on the protein of the flour and methionine were analyzed microbiologically (Hutchinson and Martin 1970). The 71 % extrac­ and the results are presented in Table 89. It was tion rate flour was milled from a No. 2 Northern noted that the variations were slightly higher when Manitoba wheat, in a Buhler laboratory mill. expressed as percent of flour than as percent of Diets in which bran replaced 0, 25, 45 and 65% of protein. In the commercial flours the correlation the white flour were compared. As the proportion coefficients between protein, threonine and of bran increased, the nitrogen level in the diet methionine were positive and significant. There increased as did (a) the growth rate per rat per day, was a significant positive correlation (r = 0.695) (b) the food intake and (c) the growth rate per rat between protein and lysine as percent flour but the per nitrogen intake. However, the growth rates correlation was negligible between protein and were still poor and well below those attainable if lysine expressed as percent protein. The average flour, without bran, is supplemented with lysine overall loss of lysine during milling was 36.3%; and threonine as found by Hutchinson et al. losses in methionine and threonine as a con­ (1956, 1964). Fine grinding of the bran did not sequence of milling appeared to be substantially affect growth rate nor food intake significantly lower. but slightly depressed the apparent digestibility of Air Classified Wheat Flour Hutchinson et the nitrogen in the diet. al. (1959) determined the amino acid composition Girish et al. (1971) reported on the protein (ion exchange chromatography) of an air classified content of 450 samples of wheat and wheat high-protein flour (Jones et al. 1959) and of its products including maida (commercial white parent flour. The results given in Table 90 are flour) and resultant atta (used in chapatis). Not compared with a typical United Kingdom baker's surprisingly, the maida, being lower in bran, flour. The amino acid composition of the air contained less protein (Kjeldahl N x 5.7) than classified flour differed little from that of the resultant atta and whole wheat. In several parent flour. instances, resultant atta was higher in protein Air classified flours of protein content as high than the whole wheat. No information is given on as 26% (Kjeldahl N x 5.7) were used in rat feeding extraction rates but ash contents in the whole trials in the form of bread by Hutchinson et al. wheat ranged from 1.3 to 1.69%. in the maida ( 1959). The bread was made in the laboratory with from 0.46 to 0.61 % and in the resultant atta from 15 g yeast per 800 g flour. Neither salt nor milk 1.26 to 1.54%. powder was included in the bread doughs but the HULSE/LAING: PROTEINS OF TRITICALE 119

TABLE 89. Protein. lysine. threonine and methionine contents (with range in parentheses) of 33 samples of Indian commercial flours. and of a composite grist wheat and laboratory-milled flour (14% moisture basis) (data from Tara et al. 1971 ).

Commercial Composite Laboratory- flour grist wheat milled flour

Protein (N x 5. 7). mean °/0 9.55 9.90 9.0 (7. 7-12. 1) Lysine g/100 g flour 0.176 0.286 0.189 (0.122-0.256) g/ 100 g protein 1.84 2.89 2.10 (l .46--2.45) Threonine g/100 g flour 0.301 0.333 0.310 (0. 252-0. 376) g/100 g protein 3 .13 3.37 3.40 (2. 71-3.3I) Methionine g/100 g flour 0.170 0.180 0. 177 (0. 129-0. 196) g/100 g protein 1. 78 1. 82 1.96 (l .53-1.98) diets as fed over 28 days from weaning contained TABLE 90. Amino acid composition of air classified added salt, a vitamin mixture and other non­ flour, its parent flour and a baker's flour (protein protein nutrients. As the level of bread protein Kieldahl N x 5.7) (data from Hutchinson et al 1959). in the diet increased from (A) 10.8% to (B) 15.0'/o to (C) 18.8% to (D) 24.6%, a moderate increase in Amino acid nitrogen rat growth was recorded. Throughout lysine (g/ 100 g total N) remained the limiting amino acid. Derived A typical Graham et al. (1969) reported that white flour high- UK• air classified to contain 21 % protein provided Parent protein baker's infant diets similar in their components to control flour flour flour casein diets. Table 91 shows the essential amino (11.3% (26.8% (12.9% acid composition (methods of determination not Amino acid protein) protein) protein) stated) of the protein of (a) human breast milk (HBM) and (b) the air classified wheat flour used, Alanine 2.70 2.87 2.87 expressed as grams per I 00 g protein and as Arginine 6. 72 2.80 2.60 2.66 milligrams per gram total essential amino acids Aspartic acid Cystine 1. 65 1 .68 (EAA). The subjects, six severely undernourished Glutamic acid 18.66 19.69 19.94 infants in the British American Hospital, Lima, Glycine 3.84 3.93 Peru, received the four experimental wheat diets Histidine 3.26 3.66 3.59 in random sequence, for 15 to 36 days each, with Isoleucine 2.57 2.56 2.55 intervening 9-day periods on casein. The four Leucine 4. 72 4.76 4.81 wheat diets, in which wheat provided the only Lysine 2.30 2.38 2.36 protein source, were fed isonitrogenously and Methionine 0.98 0.98 1.05 isocalorically per unit of body weight and in­ Phenylalanine 2.61 2.75 2.80 cluded: (A) white wheat flour (21 % protein); (B) Proline 8.30 8.75 8.83 3.89 white wheat flour plus L-lysine HCI to bring the Serine 3.90 4.08 Threonine 1. 98 2.07 2.05 lysine and threonine levels to approximately 85% Tryptophan 0.98 of the levels in HBM (i.e. equivalent to an enrich­ Tyrosine 1.41 1.46 1.61 ment level of0.12% lysine in ordinary white flour); Valine 3. 18 3 .13 3.28 (C) white wheat flour with L-lysine HCI to bring lysine to the same proportion of the essential •values from McDermott and Pace (1957). 120 MONOGRAPH IDRC-02Ie

TABLE 91. Essential amino acid (EAA) composition of human breast milk (HBM) protein and of air classified wheat flour protein expressed as grams/ 100 g protein (%protein) and asmg/g total essential amino acids (mg/g EAA (data from Graham et al. 1969).

HBM protein Wheat protein W/HBM. (%protein) (mg/g EAA) (%protein) (mg/g EAA) (%)'

Isoleucine 6.4 131 4.22 125 95 Leucine 8.9 183 6.51 193 105 Lysine 6.3 129 2.67 79 61 Phenylalanine + Tyrosine IO. I 207 8.25 244 118 Cystine + Methionine 4.3 88 3.62 107 121 Threonine 4.6 94 2.8 83 88 Tryptophan I. 6 33 I. 2 36 109 Valine 6.6 135 4.5 133 98 Total 48.8 1,000 33.77 1,000

'The~~ EAA for each amino acid in wheat as a percentage of that in HBM. amino acids, 129 mg lysine per gram EAA, as is The protein contents of (a) a hard red wheat, found in HBM protein, (see Table 91, i.e. equiva­ and (b) the 72% extraction flour milled from (a), lent to an enrichment level of 0.2% lysine in and of (c) a soft wheat, and (d) patent flour, (e) a ordinary white flour), and (D) wheat flour with straight flour and (j) a cut-off (air classified) flour L-lysine HCl to bring lysine to the same propor­ milled from (c) are given in Table 92. The essential tion of the wheat flour protein (6.3%) as is found amino acid (EAA) compositions of (a) and (c) in in HBM protein (see Table 91, i.e. equivalent to milligrams per 100 g dry weight are also quoted in an enrichment level of 0.4% lysine in ordinary Table 92 with the percentage of wheat EAA found white flour). in the fractions (b), (d), (e), and (j). The highest All the diets were well accepted and tolerated. losses on milling occur in lysine, tryptophan and Diet (B) increased rates of weight gain and nitrogen threonine. retention, stability of serum albumin and eleva­ In their review of milling principles Ziegler tion of the ratio of plasma lysine. Diets (C) and and Greer (1971) point out that the yields and (D) produced further progressive increases in protein contents of the fractions obtained by air weight gain, nitrogen retention and plasma lysine. classifying wheat flour vary with different wheats. Graham et al. (1969) conclude that enrichment The yield of the fine fraction from hard wheat of ordinary white wheat flour with lysine to the flour is much lower than that from soft wheat. The 0.12% and possibly the 0.2% level would be ratio between the protein contents of the fine beneficial in those areas where wheat provides the fraction and that of the parent flour is about main source of protein in the diet, particularly 1.5: 1 from hard wheat, 2: 1 from soft wheat. The that of infants and children. yield of medium fraction is lower, and of coarse The prolonged feeding of wheat flour enriched fraction higher, from hard wheats than from soft at the same levels as in the previous study confirmed wheats; the protein content of the medium frac­ the apparent superiority of the 0.2% level over the tion of hard wheats is about two-thirds, and of 0.12% level, and in the one case in which it was soft wheats less than half, the protein content of used, of the 0.4% level over the 0.2% level (Graham the original flour. 1971). Amino Acid Availability The effect of ex- Toepfer et al. (1969, 1972) have reported on the traction rate on the availability of the amino acids amino acid composition of selected wheats, the (assayed microbiologically) in wheat was reported flours milled from them and the products into by Calhoun et al. (1960) for lysine, and by Hepburn which they were made. The methods by which the et al. (1966) for methionine, threonine, tryptophan, amino acids were analyzed are not given but may valine, phenylalanine, leucine and isoleucine. The be expected to be published in a paper in prepara­ blend, consisting of a hard red spring wheat and a tion by Hepburn. hard red winter wheat, had a nitrogen content of HULSE;LAING: PROTEINS OF TRITICALE 121

TABLE 92. Protein content of (a) hard red wheat and (b) 72% extraction flour milled from (a); (c) soft wheat and

(d) patent flour about 54'.i0 extraction. (e) straight grade flour about 72% extraction and(/) cut-off (air classified) flour milled from (c). with essential amino acids (EAA) composition of (a) and (c) and percentage of wheat EAA found in (b). (d). (e) and (/1 (data from Toepfer et al. 1972).

(/) (e) Cut-off (a) (b) (c) (d) Straight flour Hard red 72% Soft Patent grade (air wheat extraction wheat flour flour classified)

Protein (N x 5.83 0 dry basis). 10 14.8 13.7 11. 7 9.7 10.6 11. 3

0 (" ( Essential (dry 1reigh1 0 of »·heal (dry »·eigh1 10 of \\'heal (%of wheal (% ofll'hea1 amino acids mg/JOO g) EAA) mg/ JOO g) EAA) EAA) EAA) Lysine 433 69 360 66 74 83 Cystine 310 94 223 96 101 115 Valine 742 90 581 84 89 102 Methionine 248 103 206 89 95 Ill lsoleucine 617 98 464 90 96 107 Leucine 1.068 97 812 91 95 107 Tyrosine 461 94 344 90 95 109 Phenylalanine 765 101 567 91 97 110 Tryptophan 266 83 225 68 70 77 Threonine 471 86 393 76 81 93

2.46% (Kjeldahl-Gunning procedure) and the 72% essentially similar. As in the lysine study (Calhoun extraction patent flour commercially milled from et al. 1960), the best agreement between basal this blend, a nitrogen content of2.31%. diets and between sample levels was obtained The availability of lysine was evaluated by from the relation between increase in carcass weanling rats (Calhoun et al. 1960), fed two basal nitrogen and the amount of available amino acid diets, of which wheat or flour constituted 70% by consumed. The increase in total carcass nitrogen weight. Diet (A) contained 20% wheat gluten, content was found to vary directly with the amount Diet (B) an amino acid mixture equivalent to the of available amino acid consumed when that amount of gluten in Diet (A) but from which amino acid was limiting in the diet. The avail­ lysine was omitted. Performance was rated by the abilities as calculated are shown in Table 93. gain in live weight, gain in empty body weight Saunders and Kohler (1972) have described an after removal of the intestinal contents, and gain in vitro method for the measurement of protein in carcass nitrogen over a 3-week period. The digestibility using Streptomyces griseus protease response of the samples was referred to standard (Pronase B grade, Calbiochem, Los Angeles), curves and the results compared with those followed by treatment with chick pancreas acetone obtained by microbiological assay. The closest powder. This method correlated well with an in agreement between the basal diets was found when vivo method using rats, fed isocaloric and carcass nitrogen gain was related to total available isonitrogenous diets equivalent to 10% protein, lysine consumed. On that basis the availability of the comparative standard being casein. Table 94 lysine in wheat was 75% and in the flour 72% with gives the protein digestibility determined by the the gluten basal diet, and 78% and 80% respec­ two methods on wheat mill feeds from a sample of tively with the amino acid basal diet. hard red spring wheat and also quotes the values For the other amino acids (Hepburn et al. obtained by the in vitro procedures of Booth and 1966) the wheat and flour were from the same Moran (1946), Chick et al. (1947) and Akeson and lot as in the lysine study and the methods were Stahmann (1964). 122 MONOGRAPH IDRC-021e

TABLE 93. Availability of seven amino acids in wheat TABLE 95. Extraction rate and protein content of and flour fed at two diet levels (calculated by increase in seven milling fractions of a Polish rye (data from rat carcass nitrogen) (data from Hepburn et al. 1966). Bartnik 1964).

Wheat Flour Protein content (N x 5.83 dry matter Diet Avail- Diet Avail- basis) level ability level ability Extraction (%) (%) (%) (%) rate Total Digestible' Flour (%) (%) (%) Methionine 22.5 96 25 96 45 88 50 95 Light rye flour 45.4 6.65 4.96 Tryptophan 25 88 25 88 Rye flour 60.0 6.70 4.97 40 91 40 86 Rye flour 70.0 7.29 5.26 Isoleucine 20 73 20 91 Rye flour 78. 5 8.92 6.20 35 91 35 86 Rye flour 82.0 8.63 6.00 Phenylalanine 15 76 15 90 Army rye flour 87.0 9.56 6.89 25 73 25 79 Coarse rye flour 98.0 9.79 6.64 Valine 25 76 25 78 40 72 40 82 'Bartnik (1960). Leucine 25 94 25 75 40 91 40 97 protein (dried whole milk, pulverized dried beef Threonine 30 90 30 102 and dried whole egg). 45 IOI 45 88 The growth rate method of Osborne et al. ( 1919) was used with groups of 12 rats, six male and six female of initial weight about 49 g, over six weeks. Rye Flour The composition of the seven fractions reported by Bartnik ( 1964) reported on feeding trials with Bartnik (1964) is given in Table 95. rats bred at the Institute for Food and Nutrition, When these seven fractions of rye flour were Warsaw, on seven flour fractions from a single incorporated on a 1 : 1 basis with the basal diet, sample of rye grown in Poland. The fractions the protein content in the diets rose progressively, were fed, contained in the proportion of 1 : 1, with from 9.74% (N x 6.25) to 11.08%; the proportion a basal diet simulating the average composition of of flour protein in the total protein of the diet rose, the Polish diet. The control diet was the basal diet from 37.1 % to 46.1 %. but the protein efficiency alone which provided 480 kcal and 11 .7 g protein ratio of the various diets did not differ. The weight (N x 6.25) per 100 g diet. Of this 11.7 g protein increases were approximately 2 g per gram total about two-thirds was in the form of animal protein and 2.6 g per gram digestible protein in all

TABLE 94. Protein digestibility in wheat mill feeds by in vivo and in vitro procedures (data from Saunders and Kohler 1972).

Protein digestibility,(%)

Booth & Chick Akeson & Saunders & Kohler (1972) Moran et al. Stahmann (1946) (1947) (1964) (In t•fro) (In ritro) (In ritro) (In ritro) (In ritro)

Casein control 99.3 98.5 Patent flour 93.2 96.7 99.4 99.0 98.9 Red dog 84.6 89.5 95.2 94.4 93.9 Germ 86.7 85.5 91. 5 92.8 88.3 Shorts 77.4 79. I 82.7 84.1 80.8 Bran 73.0 71. 5 74.5 72.0 69.7 HULSE;LAING: PROTEINS OF TRITICALE 123

TABLE 96. Amino acid composition of rye grain. rye flour and bran (data from Rukosuev and Silant'eva 1972).

Rye grain Sieved flour Break flour Bran Total Total Total Total protein (g/ JOO g protein (g/ JOO g protein (g/JOO g protein (g/ JOO g ('./o) product) (%) product) (%) product) (%) product)

Lysine 4.23 0.46 3.30 0.32 3.46 0. 37 4.06 0.67 Histidine 2.09 0.22 1.90 0.19 I .82 0.18 2. 19 0.35 Arginine 5.62 0.60 4.57 0.44 4.94 0.53 6.31 I .02 Aspa rtic acid 7 .16 0. 77 6. 12 0. 58 7 .16 0.80 7.47 1.22 Threonine 3. 11 0.34 2.54 0.25 3.29 0.35 3.34 0. 57 Serine 4.54 0.48 4. 56 0.44 5.05 0.55 4.53 0.73 Glutamic acid 29.91 3.25 34.46 3.22 34.94 4.03 27 .93 4. 53 Pro line 5.20 0.55 5.90 0.56 5.96 0.65 4.93 0.79 Glycine 4.79 0.52 3.73 0. 36 4.84 0. 52 5.44 0.89 Alanine 5 .13 0.55 4.22 0.41 4.78 0.51 5.36 0.88 Cystine 1.19 0. 13 I. 20 0. 13 0.91 0.08 1.90 0.32 Valine 5.56 0.60 4.92 0.48 5.45 0.60 5.32 0.86 Methionine 0.65 0.07 0.21 0. 02 1.08 0.10 0.44 0.08 lsoleucine 3.88 0.42 3. IO 0. 30 4.09 0.44 3.69 0.61 Leucine 7.00 0.75 5.85 0.56 7.33 0.81 6.76 1.09 Tyrosine 2.86 0.31 2.67 0.26 2.90 0.31 2.66 0.43 Phenylalanine 5.25 0.56 4.99 0.48 5.57 0.62 4. 56 0.73 Ammonia 2.92 0. 30 2.74 0.28 2. 16 0.22 2. 19 0. 37 the diets. The total weight increases over the six in feeding tests with rats using the original growth weeks varied from 75 g to 96 g but in parallel with method of Osborne et al. (l 919)and a modification. the percentage protein of the flour added to the The diets were fed at 5.5% protein (N x 6.25) basal diet. The weight increases of the animals on level. At this low level of protein, both rye flours the test diets per gram protein consumed was gave very similar biological values by both higher in the test diets than on the basal diet alone. methods, means of 45.8% and 45.9% for the 98% Bartnik (1964) comments that these results could extraction flour and 44.8% and 49.0% for the 45% indicate, so far as growth rate is concerned, either extraction flour. These values were approximately that there is no difference between the protein of that of whole egg. the different milling fractions of rye flour examined Szkilladziowa ( l 960a) determined the biological or that the protein in the basal diet balanced the value of rye flour of three extraction rates, (A) protein in the rye flours. He considers that the 98%, (B) 87% and (C) 45% by the rat growth probability is that there is no difference between method of Osborne et al. (1919) and by the the proteins of the milling fractions. This view is chemical method of Block and Mitchell (1946). supported by the findings of Szkilladziowa ( 1961 ), Wheat flour of (D) 98% and (E) 50% extraction who used rats from the same Institute strain as was also included, egg providing the control diet. Bartnik (1964). The average level at which the protein (N x 6.25) The digestibility coefficients of these seven was fed varied, 8.3% in Diet (A) (98% extraction fractions with three other rye flour fractions and rye), 8.5% in Diet (B) (87% extraction rye), 5.3% four of bran were reported by Bartnik (1960). in Diet (C) (45% extraction rye), 8.7% in Diet (D) Digestibility decreased with increasing extraction (98% extraction wheat), 8.8% in Diet (E) (50% rate. However, since the protein content rose with extraction wheat). extraction rate, there was a net increase in The biological values for the rye flours (Osborne digestible protein from the flours of higher et al. 1919) were 54 (Diet A), 52 (Diet B) and 30 extraction rates. (Diet C) respectively, and for the wheat flours 47 Szkilladziowa (1961) compared Polish rye flour (Diet D) and 23 (Diet E). The equivalent values by of 98% extraction and of 45% extraction with egg the chemical method of Block and Mitchell 124 MONOGRAPH IDRC-02le

(1946) were (A) 41, (B) 39 and (C) 37, (D) 40 and from a mixed grist of Spanish grown wheats (see (E) 35. Table 84). Baked alone, the triticale flours gave Rye flours of similar extraction rates, (A) 98%, poor bread compared with the wheat flour control (B) 87% and (C) 45%, and wheat flour of (D) 98% but mixtures of 20 parts triticale flour and 80 parts and (E) 50% extraction rates were also examined wheat flour produced bread superior in overall by Szkilladziowa (I 960b) in feeding trials with 290 baking quality to the wheat flour control, probably white rats using the method of Miller and Bender because the triticale corrected the deficiency in (1955). Various levels of protein (N x 6.25) were diastatic activity which is general in Spanish fed (between approximately 5% and 9% in the wheat flours. Vallejo et al. suggest that the addi­ diets) but the mean NPU values were respectively tion of triticale flour would improve the nutritional (A) 55.9, (B) 58.8 and (C) 61.0 for the rye flours quality of bread protein. and (D) 52.9 and (E) 43.6 for the wheat flours. The spring triticales 6T A204 and 6T A206 and Kurzepa et al. (1960) reported on the essential the winter triticales TR385 and and TR386 were amino acid content (assayed microbiologically) of compared with two winter wheats, Scout and IO flours of differing extraction rates and four Lancer, and two spring wheats Inia Res. and Chris. brans from a single sample of experimentally for the production of bread and rolls (Lorenz milled Polish rye. As the extraction rate rose from 1972). Characteristics of the flours are given in 45.5% to 98%, the content of lysine and arginine Table 97. The values, where they may be com­ in relation to total nitrogen increased and the pared, are similar to those given by Lorenz and content of leucine decreased. The content of the Maga ( 1972) (see Table 76). The triticale bread had other amino acids determined: phenylalanine, a mild rye flavour. Some changes in procedure tryptophan, histidine, valine, isoleucine, methio­ were needed to produce the best results. nine and threonine did not alter substantially. Lorenz et al. (I 972b) discuss more fully the Rukosuev and Silvant'eva (1972) compared the mixing and baking properties of these triticale amino acid composition of the proteins in a varieties. The winter triticales produced bread sample of whole rye, 1966 harvest, sieved rye of lower specific volume, more open grain, slightly flour (63% extraction), break flour (87% extrac­ harsher texture, and slightly darker colour than tion) and bran. The amino acids, determined by the winter wheats. The spring triticales produced autoanalyzer, are quoted in Table 96. bread of acceptable quality, 6TA204 had the highest volume of all the breads baked and 6T A206 was only slightly inferior to the spring Secondary Processing winter wheat bread. Bushuk (1972) reported that acceptable bread Triticale may be obtained from triticale flour using a No published work has so far been encountered mechanical development process. on the effects of processing for human consump­ Tsen et al. (1973) compared the baking quality tion on the nutritional value of triticale-containing of three hexaploid triticales of protein (N x 5.7, products. The literature covering such effects on 14% moisture basis) of(!) 11.1%, (II) 13.6% and the nutritional value of wheat and rye is con­ (III) 12.8%, and a wheat flour of 12. 7%. The siderable and is referred to later in this chapter. amino acid composition (by autoanalyzer) of Papers have, however, appeared on the utiliza­ triticales I and II and the wheat flour are given in tion of triticale in baked and other products, and Table 98. Acceptable bread could be obtained these are referred to briefly below. from the triticales by eliminating bulk fermentation Bread Unrau and Jenkins (1964) conducted and, for triticales II and III, adding 0.5% sodium baking tests using the hexaploid triticale samples stearoyl-2 lactylate, 0.25% sucrose tallowate, or described above under Milling (see Table 74 and 0.25% ethoxylated monoglyceride. Triticale I 75). Baked alone, the triticale did not give good required the addition of 20% wheat flour with results but when blended with Pembina wheat 0.5% sodium stearoyl-2 lactylate, or 60% wheat flour, up to a limit of 40% triticale to 60% Pembina, flour alone. Triticale bread staled twice as rapidly loaf volumes were comparable to those when as wheat bread, as measured by changes in crumb Pembina was baked alone. firmness with a Bloom Gelometer. Vallejo et al. (1969) compared experimentally Cake Thompson ( 1971) and Thompson and milled triticale flours with a similarly milled flour Vaisey (1971) cited by Bushuk (1972) found that HULSE/LAING: PROTEINS OF TRITICALE 125

TABLE 97. Characteristics of wheat and triticale cultivars (14% mois/Ure basis: yield lhrough JOO mesh screen) (data from Lorenz 1972).

1970 crop 1971 crop

grain flour grain flour protein protein flour flour protein protein flour flour (N x 6.25) (N x 5.7) ash yield (N x 6.25) (N x 5.7) ash yield ('/o) ('/o) ('/o) (%) (%) (%) ('/o) (%)

Spring triticale 6TA204 14.6 11. 3 0.53 42.1 15.7 12.7 0.54 61. 9 6TA206 14.5 11. 7 0.54 42.8 15.2 12.8 0.53 61. 5

Winter triticale TR385 12.0 8.7 0.47 33.9 13. I 11.6 0.49 56.4 TR386 10.8 8.6 0.47 32.7 13.4 11. 5 0.46 58.2

Spring Wheat Inia Res. 12.7 12.4 0.46 45.2 Chris 14. 3 14.0 0.46 69.1

Winter wheat Scout 12.0 11.6 0.43 57.0 12.5 11. 7 0.44 66.8 (No details are given for Lancer)

cake made from chlorinated triticale flour, even TABLE 98. Amino acid composition of triticale and at the highest levels of chlorination examined, was wheat flours in grams amino acid per 100 g Kjeldahl inferior to that obtained from standard cake flour, protein (data from Tsen et al. 1973). as measured by volume, subjective crumb scores, textureometer evaluation of hardness, cohesive­ Amino Acids Triticale I Triticale II Wheat ness, gumminess and adhesiveness of the cake 2.85 crumb. Lysine 2.63 2. 16 Histidine 2.03 2.23 2.24 Alimentary Pastes Lorenz et al. (l 972a) Ammonia 3.82 3.76 4. 33 prepared acceptable noodles from a triticale flour Arginine 4.13 4. 39 3.88 of unspecified origin, of 9. 3% moisture and 13. 3% Aspartic acid 5.73 6.45 4. 54 protein (N x 5.7). Threonine 2.98 3.36 3 .06 Bushuk (1972) reported that spaghetti made by Serine 4.70 5.25 5. 58 laboratory equipment from triticale semolina was Glutamic acid 32.03 37.22 45.85 darker and inferior overall to spaghetti from durum Proline IO. 77 12.51 12.24 wheat. On cooking, triticale spaghetti behaved Glycine 3 .62 3.91 4.09 similarly to durum spaghetti and had a rye Alanine 3.40 3.81 3.52 2.74 flavour. i cystine• 3.27 3.56 Valine 4.43 4.86 5. 17 Breakfast Cereals T riticale made into break- Methionine' 1.60 1.44 1.08 fast cereals, flakes, puffs and muffets ("shredded Isoleucine 3.37 3.83 4.26 wheat" type product) on a commercial plant were Leucine 6.76 7.45 8 .16 similar to those made from wheat but with a slight Tyrosine 3.07 3.35 4.01 rye flavour (Bushuk 1972). Phenylalanine 4.77 5. I I 5.99 Malting and Brewing Pomeranz et al. (1970) reported on the use of triticale in malting and •By oxidation. brewing. The triticales used are characterized in Table 99. Beers from the worts of 6T204 (Texas), Rosner (Winnipeg and North Dakota), 6714 and In a later paper, Pomeranz (1971 b) compared 6804 (Winnipeg) had both satisfactory clarity­ the Kjeldahl nitrogen contents and amino acid stability and gas stability. composition (Robbins and Pomeranz 1971) of the 126 MONOGRAPH IDRC-02Ie

TABLE 99. Kernel weight, and nitrogen in grain and in malt (dry matter basis) oftriticales (data from Pomeranz et al. 1970).

Kernel Kjeldahl weight nitrogen Nitrogen Selection Location (mg) (%in grain) (%in malt)

6T204 Bushland, Texas 27.6 3.20 3.33 6T208 Bushland, Texas 25.6 3.26 3.35 6T209 Bushland, Texas 27.7 3 .14 3.23 6450-3-1 Bushland, Texas 20.2 3. 10 3.24 Rosner• Winnipeg, Canada 33.3 2.48 2.48 6714. Winnipeg, Canada 30.4 2.84 2.88 6804• Winnipeg, Canada 30.3 2.65 2.76 Rosner Fargo, N. Dakota 29 .6 2.94 2.94 6437-6 Madison, Wisconsin 28.7 2.96 2.98 6450 Madison, Wisconsin 25.9 2.82 2. 93

•strains from a double cross in which two varieties of Triticum durum Desf., a variety of T. persicum, and a fourth Triticum species were used.

TABLE 100. Nitrogen contents and amino acid com­ TABLE IOI. Effect of malting on Kjeldahl nitrogen position (grams/ 100 g amino acid recorered) of rye, and lysine content of triticale, wheat and rye (data wheat and triticale grains grown under comparable from Pomeranz 1971b). conditions (data from Pomeranz 1971b). Lysine Rye Chris Rosner (g/JOOg (composite) Wheat Triticale Nitrogen amino acid (%) recovered) Kjeldahl Nitrogen,% 2.38 3.38 3.23 Rosner triticale Amino acids grain 3.23 2.6 Lysine 3. I 2.4 2.6 malt 3.35 3. I Histidine 2.0 2.2 2.0 sprouts 4.76 4.8 Ammonia 3.5 3.9 3.4 Arginine 4.9 4.1 5.0 Chris wheat Aspartic Acid 7.5 5.0 6.5 grain 3.38 2.4 Threonine 3.1 2.6 2.9 malt 3.36 2.7 Serine 4.2 4.2 4.2 sprouts 5 .10 5.7 Glutamic acid 27.2 33.2 34.2 Rye Pro line 13. I 10.8 10.6 grain Cystine/2 0.3 1.0 0.5 2.38 3. I malt 2.36 Glycine 4.1 3.8 3.8 3.9 4.50 Alanine 4.1 3.3 3.8 sprouts 4.8 Valine 4.5 4.2 4.1 Methionine 2.3 2.1 I. 8 Jsoleucine 3.4 3.5 3. I Table 100. When malted under comparable Leucine 5.7 6.4 5.4 conditions all three cereals showed a decrease in Tyrosine 2.2 2.6 2.3 glutamic acid and increases in lysine, arginine, Phenylalanine 4.7 4.7 4.0 aspartic acid, valine, isoleucine and leucine. The increase in nitrogen and lysine in malts and malt sprouts is shown in Table 101. grain, malt and sprouts of Rosner triticale, Chris W. 0. S. Meredith (1969 unpublished report) wheat and a composite rye, all grown in Bushland, cited by Bushuk (1972) malted triticale to produce a Texas, USA in 1969. The Kjeldahl nitrogen and product of high enzymatic activity which, it was amino acid composition of the grain is given in concluded, could be used successfully in brewing. HULSE/LAING: PROTEINS OF TRITICALE 127

TABLE 102. Nitrogen and protein content (N x 5.7, dry matter basis) of Chilean wheat and wheat products, and their NPU0 ., percentage of protein calories in each diet (P), NPU,, and net protein concentration (NPC =

NPU0 P x P/100) (data from Ballester et al. 1962).

Total Total N protein p (%) (%) NPU0 " NPU,. NPC

Wheat I. 75 9.98 40.2 10.0 40.3 4.0 "Mote" of wheat I. 75 9.98 31.3 10.0 31.3 3.1 Flour (76% extraction rate) I. 50 8.50 38.6 8.4 38.6 3.2 Bread 1.65 9.40 47.9 9.4 49.3 4.5 Roasted flour 1.44 8.21 34.1 8.1 34.1 2.8

Finney et al. (1971) cited by Bushuk (1972), bread was about 15% less than that of the flour found that triticale malts were superior to barley from which it was baked. malts in breadmaking. Burke (1960) determined by autoanalyzer the Miscellaneous Products Bushuk (1972) cites lysine, histidine and arginine contents of bread E. N. Larter (1972 unpublished report) as report­ made from 90% extraction (South African) flour ing the test marketing of triticale pancake flour and and of the unbaked dough. A loss of about I 0% of a mix based on triticale flour for pizza pastry. lysine occurred during baking but histidine and arginine were not noticeably affected. Wheat and Rye Ballester et al. (1962) reported on the nutritive Wheaten Bread Hepburn et al. (1957) as- value of Chilean cleaned, milled white wheat, the sayed, microbiologically, the amino acids in the "mote" prepared from it, white flour available crust and crumb of bread from wheat flour and commercially in Santiago (76% extraction), pan compared the results with those from the whole bread and roasted flour. The "mote" was pre­ loaf. Only in the case of lysine was the value for pared by boiling (for about 2 hours) and decorti­ the crust significantly less than that for the crumb. cating the wheat in an alkaline lye, grinding and The content of lysine in the whole bread sample washing with water. The fresh product contained equalled the value expected from the proportion about 65% water. It was dried at temperatures of crust and crumb in the bread. below 60°C and ground. The bread was prepared McDermott and Pace (1957) determined colori­ with flour 100 g, salt 3 g, sugar 3 g, yeast 3 g and metrically the amino acid contents ofhydrolysates cooking fat 3 g and was baked at 220°C for 20 to of wheat flour, and of the whole bread and of 22 min. The roasted flour was made from whole, bread crust, made from the flour. The contents of cleaned, white wheat, roasted at 160°C. phenylalanine, tryosine and serine in the bread The analysis of the products and results of hydrolysate were lower by between 5 and 7% NPUs (Miller and Bender 1955; Miller and Payne than in the flour hydrolysate. The contents of 1961) are given in Table 102. phenylalanine, tyrosine and lysine were 15 to 16% The NPU" of the bread was higher than the lower in the crust than in the flour hydrolysate. NPU" of the wheat and of the flour. The roasted Using microbiological assay methods, Horn flour did not support growth and by the tenth day et al. (1958) found that no destruction of amino the rats on that diet had fallen below their starting acids occurred during fermentation of bread weight. The best rate of growth was shown by the doughs prior to baking but losses of cystine, lysine rats on the wheat diet, followed by those on the and methionine during baking were statistically bread, commercial flour and "mote" in that order. significant in small experimental loaves in which Gotthold and Kennedy (1964) examined the the ratio of crust to crumb is greater than in larger effect of baking and steaming on the ingredients of commercial loaves. whole wheat bread. The same bread formula Clegg and Davies (1958) using a chemical containing whole wheat flour, protein content method (Bruno and Carpenter 1957) found that 15.5% (Kjeldahl N x 5.83) was baked and steam­ the apparent available lysine in the crust of wheat cooked and the nutritional value of each compared 128 MONOGRAPH IDRC-02le with that of the unbaked ingredients by the follow­ and (b) a weak flour by both CBP and a con­ ing four criteria derived from rat feeding experi­ ventional breadmaking method were compared ments: (a) change in body weight, (b) PER and no significant difference was recorded between (modified method of Osborne et al. 1919), (c) BV methods for either of the two flours. (N-balance method of Mitchell (1923-24) and Rye Bread Kon and Mark use (1931) report Mitchell and Carman (1926) (baked bread only), feeding commercially baked rye bread from 70% and (d) NPU (Bender and Miller 1953; Miller and extraction flour as 95% of the diet of female rats. Bender 1955). The crust and the crumb of the same bread were By all methods, after 28 days, there was little similarly fed to male rats. The PER for the whole significant difference between the unbaked in. bread, the crust and the crumb are given below: gredients and the steamed bread whereas the Protein baked bread showed significantly lower values. (N x 6.25) The following are percentage differences for each (%) PER of the method: Whole bread (female rats) 6.4 I. 25 Difference between Crumb (male rats) 6.7 I. 26 unbaked ingredients Crust (male rats) 6.3 0.83 and baked bread (%) Body weight 53 Although the rats in the trials were of different BV 8.6 sexes, and the results are therefore not strictly PER 11.0 comparable, the findings indicate that, as reported NPU 15.0 elsewhere, there is a significant loss in protein value in the crust, but the difference between The above were all computed after 28 days' whole bread and bread crumb is not of con­ feeding tests. After 10 days the results were sequence. inconclusive. Kofranyi (1957) reported on nitrogen balance While all the 28-day results reveal the same tests, conducted on himself, to compare the general trend, the difference in magnitude among biological value of milk protein with rye protein. methods is worthy of note. In all, the tests were conducted in stages over a Gotthold and Kennedy (1964) cite Adolph and period of 10 months. The rye protein was taken in Tsui (1935) who compared the growth-promoting the form of rye bread of0.84 to 0.91 % N, the only qualities of steamed and baked white bread. They "foreign" protein amounting to 0.074 g N per day. fed the two breads ad libitum to weanling rats for He found that the addition of glutamic acid did 8 weeks and reported satisfactory growth and not improve the biological value of the rye protein little, if any, difference, between the two types. and slightly reduced that of the milk protein. The Gotthold and Kennedy (1964) from an analysis of values found in the nitrogen balance tests were so the growth curves given by Adolph and Tsui low that absolute figures for biological value were (1935) concluded that the rats fed baked bread not obtained. He concluded, however, that pro­ gained 28% less in 4 weeks and 15% less in 8 portionally the biological value of rye protein weeks than did those fed steamed bread. This was 22% lower than that of milk protein. difference is in agreement with the findings of Kofranji and Muller-Wecker ( 1960) reported on Gotthold and Kennedy in which the rats fed dietary studies with three men and one woman to baked bread gained 27% less weight in 4 weeks compare the biological values of the proteins of than those fed steamed bread. milk, egg, rye, and wheat. They report that the The Chorleywood Bread Process (CBP) is a ratios were respectively 100: 100:83:41. When method of making bread from wheat flour in lysine was added to the wheat and rye diet, the which the normal 2 to 4-hour period of bulk biological values improved; the addition of fermentation of the dough, customarily used in the methionine also effected some improvement but United Kingdom, is replaced by a few minutes of the addition of leucine did not. intense mechanical agitation. The effect of this The minimum protein requirements of the process on the nutritional quality of the flour individuals were found to differ considerably but protein was examined by Chamberlain et al. ( 1966). reproducible results were obtained, within indi­ The NPUs of bread made from (a) a strong flour viduals and within protein sources, provided the HULSE/LAING: PROTEINS OF TRITICALE 129 results of the first I 0 days of the experimental Microwave baking Clarke and Kennedy period were not included. No agreement was (1962) evaluated the effect of baking, by micro­ found between the results of the feeding experi­ wave and in a conventional oven, on the protein ments and biological values calculated from amino quality and lysine availability of whole wheat acid analyses whether by the chemical score bread with an addition of milk powder. The method of Mitchell and Block (1946) or with the protein quality was assessed by gain in body EAA index of Oser (1951). weight, PER, and increase in carcass nitrogen of Crisp Bread Kihlberg and Ericson (1964) rats, and apparent lysine availability by faecal in rat feeding trials found marked differences excretion. The diets provided approximately 10% between a rye flour composed of 70% Russian and protein. The dough was baked in an air-oven in 30% Argentinian rye and the corresponding metal pans and by microwave baking in glass crisp bread. Nitrogen in the diet was 1.63% and containers. the differences are shown below: The nutritional value of the air-oven baked Nitrogen bread was significantly lower than that of the efficiency unbaked ingredients by all the criteria used, the Average ratio decreases ranging from 24 to 45%, and between weight (g wt. gain/g N 20 and 32% lower than that of the microwave baked (gain g/day) eaten) bread. The body weight gains of the animals fed the microwave bread diet were significantly lower Rye flour 1.99±0.11 12.68±0. 35 than those fed the unbaked ingredients, but since 7.73±0.47 Rye crisp bread 1.15±0.09 they ate less food, the gain per unit of protein During baking, crisp bread suffered a much eaten was not significantly different. The apparent greater decrease in protein value than did soft availability of lysine was about 20 to 30% less in bread probably because of the higher average oven baked bread than in the unbaked ingredients temperature reached. As reported elsewhere, but little difference was found between the lysine loss is much greater in the crust than the ingredients and the microwave baked bread. crumb of "soft" breads. Wheat crisp bread fed Chapatis Milner and Carpenter (1969) pre- at the same time did not even maintain the starting pared chapatis from three parts whole wheat flour weight of the young rats. and one part water, rolled into thin discs and Munck (1972) described experiments with cooked on a hot plate at 210°C for 2 min each Swedish whole wheat bread baked in thin flat side, without fat. After air drying, they were cakes containing 54.3% water in the dough. For ground for incorporation in a diet for male and (A) soft bread, the dough was baked at tempera­ female weanling rats. The chapatis, representing tures of 450--340-400°C in sequence over 2.0 min approximately 60 to 64% of the diet, were fed to and for (B) hard bread, at 430--340--380°C in provide I 0% crude protein (N x 6.25). sequence over 3.6 min. The moisture content in the The PER values were higher for chapatis than cooled bread was for (A) 26.3% and for (B) 11.0%. for uncooked wheat. Milner and Carpenter ( 1969) The composition of lysine, histidine, arginine, consider that this improvement is due to increased threonine and methionine in the soft bread palatability of the cooked product and hence to changed little in comparison with the whole wheat increased weight gain rather than to any improve­ meal, but in the hard bread all the amino acids ment in the nutritional value of the protein showed a reduction, especially marked in the case resulting from the processing. of lysine. Chapatis were also included at a 13% protein In mice feeding trials, the whole wheat meal and level in a diet for chicks. The responses to the the breads were fed at about 14.0% protein (dry chapati diet and to the control, untreated wheat, matter basis). There was little difference in gain diet were similar. per animal, food consumption and grams weight Shyamala and Kennedy (1962) compared, in gain per gram protein consumed between the whole rat feeding tests, (A) uncooked whole wheat flour, wheat and the soft bread diets (lysine consumed 1.67% nitrogen (N), (B) chapatis of whole wheat per animal 234 and 277 mg respectively) but there flour, 1.60% N, (C) chapatis of 90% whole wheat were reductions from the hard bread diet (lysine flour and 10% defatted soya flour, 2.35% N, (D) a consumed per animal 188 mg). skim milk diet, 5.93% N. Each diet provided 1.5% 130 MONOGRAPH IDRC-021e

N and the tests continued over 4 weeks. Body centration of lysine in the standard biscuits was weight gains and PERs were determined. The 126.4 ± 9.7 mg/g N; the mean percentage of factors used to convert N into protein were 5.83 for Kjeldahl N was 2.18. The lysine content of the the whole wheat flour, 5.79 for the flour plus de­ fl our was 139. 3 mg/ g N. Therefore 9. I% of the fatted soya flour and 6.38 for the milk diet. lysine in the flour was destroyed or rendered The chapati diet was significantly better than unavailable to the microorganisms as a result of uncooked flour, which Shyamala and Kennedy baking. As baking time increased so did the loss attribute to the destruction of trypsin inhibitor(s) of lysine; 18.1% in 25 min, 27.0% in 30 min. When but which Bender (1969) suggests is the result of L-lysine HCI was addrd the proportional losses improved palatability. The soya supplemented were comparable to those of the lysine naturally chapatis produced 51 % increase in PER over present. Variations in the other ingredients had no the cha pa tis without soya and were not significantly effect except that, for any given baking time, higher different from the milk diet in body weight gain proportions of sugar increased the loss of lysine. or PER. The concentration of lysine was significantly In a second series, puris (similar to chapatis lower in the crust than in the crumb. but deep fried in hot vegetable oil) displayed PER Cake An unbaked cake mixture (with a values 22% lower than whole wheat chapatis. The contribution of essential amino acids approxi­ food intake from the puri diet was lower than that mating that of whole egg) was found by Block et al. from the chapati diet. (1946) to have a protein efficiency between 3.3 Baking Powder Biscuits (Scones) The effect and 3.5 when fed to protein-deficient adult rats. of baking on the PER of the protein in baking When the mixture was baked into a cake, and dried powder biscuits (scones), muffins and griddle­ before feeding, the efficiency fell to 2.4, and when cakes was assessed by Kennedy and Sabiston slices of the cake were toasted at I 00 to I 30°C the (1960) in rat diets providing approximately I 0% efficiency fell to 0.7. The reduction was apparently protein fed ad libitum over 4 weeks. Cookies were due to the destruction or non-availability of the also fed but assessed over 3 weeks. The PERs for lysine since the addition of 0.63% of L-lysine to the unbaked ingredients were significantly greater the toasted cake restored the protein efficiency than for the baked products. The reductions in to 3.2. PER of the baked products compared to unbaked Alimentary Pastes Cubadda et al. (1970) ingredients are shown below: conducted rat feeding trials to determine the PER effect of drying at various temperatures on the reduction nutritive value of alimentary pastes. Earlier work Baking compared by the same authors had shown that using the Baking tempera- to unbaked chemical method of Carpenter (1960) as modified time ture ingredients (Roach et al. 1967), and a microbiological method, (min) (°C) (%) there was a reduction in available lysine depending on the drying temperature, amounting to a 40% Biscuits (scones) 12 218 12.7 loss at 80°C. In the present experiments, the pasta Muffins 23 204 14.5 dried at (A) ca 20°C, (B) 60°C and (C) 80°C con­ Griddlecakes 2 204 17.8 tributed 89% of the 10% protein (N x 5.7) diet Cookies IO 177 15.5 for male and female albino rats, six per group. The PER, NPU and FCE (Food Conversion No apparent relation was noted between degree of Efficiency) values are given in Table 103. The reduction in PER and the extent of colouration lysine content of the pasta (determined by of the crust. It is not mentioned whether the autoanalyzer) fell from 1.84% of protein for approximate ratios of crust: crumb were deter­ pasta (A) to 1.75% for pasta (B), to 1.67% for mined in the various products. pasta (C). The reductions in nutritional value, Clark et al. (1959) assayed microbiologically and in lysine, with increase in drying temperature the loss of lysine in powder biscuits (scones) are significant. baked at 232°C for 20 min. Biscuits were prepared Toepfer et al. (1969, 1972) compared the amino for analysis by drying at room temperature and acid composition of a durum wheat of 15.6% extraction by petroleum ether. The mean con- protein (N x 5.83, dry basis), with that of the HULSE/LAING: PROTEINS OF TRITICALE 131

TABLE 103. Food consumption, weight gain, FCE (food conversion efficiency), PER and NPU of male and female rats fed pasta dried at various temperatures (Cubadda et al. 1970).

Average food Weight consumption gain per head Treatment (g) (g) FCE PER NPU

Males Diet(A) 19.41 ±3.29 163.7 0.116±0.017* 1.07 ±0.15 52.0 (Ambient temperature) ca 20°C Diet (8) 18 .09± 5.02 161 .8 0.109±0.014 I .OJ ±0.16 51 .6 (Temp. 60°C) Diet(C) 16.25±4.04 161 .2 0.104±0.017 0.96±0.18 46.6 (Temp. 80°C) Females Diet(A) 24.33±5.46 175.4 0.135±0.022 1.25±0.21 51. I (Ambient temperature) ca 20°C Diet (8) 19.33±4.03 168.3 0.118±0.017 1.09±0.18 47.8 (Temp. 60°C) Diet (C) 16.67*±2.29 162.5 0.108±0.010 0.98** ±0. 12 46.2

*P<0.01. **P<0.001. semolina ( 15.2% protein) and the macaroni protein foods for infants and malnourished ( 14.9% protein) prepared from it. persons, especially in developing countries. The macaroni was prepared commercially with Bulgur Bulgur or burghul consists of par- a dough absorption of 25% and a drying time of 41 boiled wheat and is a common food of the Middle hours at 38°C. Lysine (determination method to be East. In its preparation, the whole wheat is published) in the durum wheat was 433 mg/ I 00 g steeped in hot water to swell the grain, then boiled grain, dry weight. Lysine in the semolina was 76~~ and finally dried (Milner and Carpenter 1969). and in the macaroni 77% of that found in the Jamalian and Pellett ( 1968) determined the amino wheat. acid composition (by autoanalyzer, tryptophan Breakfast Cereals Griswold ( 1951) review- by Lombard and Delange 1965) for two samples ing the literature on the effect of heat on the of bulgur. The analysis for the essential amino nutritive value of proteins concluded that the acids is given in Table I 04. nutritive value of the protein of cereals was not damaged by cooking with water but was lowered TABLE I 04. Essential amino acid composition of by toasting, as in processing flaked or puffed bulgur (data from Jamalian and Pellett 1968). cereals. Bender ( 1966) in a later review concluded that heating, rolling and flaking processes do not (mg/g N) affect the protein but the process of "explosion" puffing, in which temperatures of about 200°C Tryptophan 45 and pressures of I 00 to 200 lb are used, may reduce Threonine 172 PER values significantly. For example, wheat Isoleucine 195 balls fell in PER from 1.6 to 0.3 during puffing. Leucine 390 Since breakfast cereals are not normally a major Lysine 160 Methionine 90 component of diet, and since they are usually a Cystine 123 vehicle for milk, these losses are not regarded as Phenylalanine 253 serious in developed countries. However, high Tyrosine 183 temperature extrusion and explosion puffing Valine 230 should be avoided in the processing of cereal and 132 MONOGRAPH IDRC-021e

The data of Jamalian and Pellett ( 1968) does not oven-dried at a temperature below 40°C (mild relate the nutritive value of the protein of the treatment); (C) wheat steeped at 80°C for 70 min, bulgur to that of its original wheat. The effect of cooked at I 0 lb pressure for 60 min and oven­ the processing is however reviewed and discussed dried below 40°C (moderate treatment); (D) wheat by El-Lakany et al. (1969) and by Milner and steeped at 60°C for 4 hours, cooked at 15 lb Carpenter ( 1969). pressure for 15 min, oven-dried below 40°C, then El-Lakany et al. ( 1969) reviewed earlier work on steeped at 80°C for 70 min, cooked at 10 lb pres­ the effect of various treatments on the biological sure for I hour and oven-dried below 40°C value of the protein of whole wheat. They cite (severe treatment). Beaudoin et al. (1951) who found that the bio­ The results are given below: logical value improved when the wheat was cooked PER NPU in boiling water as in the preparation of shredded (A) Raw wheat I. 12 39.7 wheat, and also cite Shammas and Adolph (1954). As percent of control (A) Yang and Al-Nouri (1962) and Kohler (1964) who (B) Mild treatment 110 10 I reported that the nutritive value of wheat is (C) Moderate treatment 89 91 unaffected by mild cooking as in the preparation (D) Severe treatment 85 90 of bulgur. The higher PER given by bulgur (B) was In their own experiments, El-Lakany et al. probably due to improved palatability; NPU was (1969) compared the metabolizable energy (ME) unaffected. for chicks of two samples of wheat of protein In further trials, growth response and protein 15.4% and 12.1% respectively (Kjeldahl N, con­ efficiency of chicks was used to compare (A), (B) version factor not stated). The wheat was: and (D) with: (E) wheat steeped in a large volume (A) untreated, ground; (B) autoclaved for 60 min of water at 80°C for 70 min, and then dried; (F) at 15 lb pressure. fan-dried at room temperature wheat steeped as for (E), boiled for 15 min in and ground; (C) water-soaked overnight, auto­ excess water, drained and oven-dried at a tempera­ claved for 60 min at 15 lb pressure, fan-dried at ture below 40°C. room temperature, ground; (D) frozen for 5 days Growth response and "PER" were significantly at -4°C, thawed and ground. lower for (D) than for (A); there was no positive All the treatments significantly improved the response to the other products. metabolizable energy (ME) content of the wheat When (A), (E) and (F) were fed at 92.7% of the for chicks compared to the untreated control diet to rats, PERs were significantly higher than wheats. Soaking the wheat in water before auto­ control, (E), 153% and (F), 145%. claving in treatment (C), did not enhance the When (A) and (E) were fed in a further experi­ effect of autoclaving. The cause of the significant ment with rats, PER and NPR increased signifi­ increase in ME values following freezing is not cantly but NPU did not: (A), 36.9; (E), 37.8. In explained. this trial, the appetite quotient (rate of food intake Wheat of protein content 15.4% was fed, (A) in relation to its metabolic size, i.e. body weight) untreated, and after autoclaving for (E) 60 min, 0.88, was significantly higher for (E) than for (A), (F) 90 min and (G) 120 min at 15 lb pressure. that is the steeped wheat was more palatable than followed by fan-drying at room temperature. the uncooked control. The digestible energy (cal/g Again, the ME value of the wheat autoclaved for dry matter) of (E) was 4% higher than for (A). 60 min was significantly improved over that of the Milner and Carpenter (1969) also compared untreated control but the ME values of treat­ PERs and appetite quotients of rats fed 13% ments (F) and (G) were significantly reduced com­ protein diets in which the sole source of protein pared to that of the untreated control. was (A) untreated wheat; (G) wheat germinated Milner and Carpenter (1969) compared the until the grains just began to sprout, dried; (H), PER and NPU values for male and female rats, (G) treated as (E); (J), (G) treated as (B); (K), (G) fed over I 0 days, on American hard winter wheat treated as (B). The trend in results was as for the of 17'/~ protein (N x 6.25, dry matter basis) other experiments; the best performance being treated as shown below: that of (H), the steeped, germinated wheat. (A) Raw wheat; (B) wheat steeped at 80°C for There was no difference in total lysine (Moore 70 min, cooked at 5 lb pressure for 30 min and et al. 1958 and Carpenter et al. 1963) between (A) HULSE/LAING: PROTEINS OF TRITICALE 133 and (D) but FDNB-available lysine (Carpenter (B), wheat from the same grist, steamed at the 1960) fell by 14% in the production of (D), mill, and laboratory-milled as (A). Flour (B) gave severely treated bulgur. Total cystine also fell by a significantly better growth rate of 4.5 compared 14% but none of the other amino acids investigated to 3.3 from flour (A). showed any change. Heat processing had no In further trials, six rats per group were fed significant effect on the availability of leucine, (C), Manitoba wheat, unsteamed; (D), the same tryptophan and methionine determined micro­ as (C), laboratory-steamed, as wholemeal, in biologically. diets containing 2.55% Kjeldahl nitrogen over In summary, mild heat processing of wheat 28 days. Wholemeal (D) gave a significantly higher improved its palatability for rats which grew growth rate, 7.45, than wholemeal (C), 6.55, faster and gave higher PER values than when fed showing that the effects were not caused by unprocessed wheat. Severe heat treatment resulted variability in the milling process. in a I 0% fall in NPU with some destruction of The improvement given by the steamed wheat cystine and reduced availability of lysine. diets were shown, statistically, to be accounted for, Pence et al. ( 1965) reported on the results when almost entirely, by the increased food intake on seven samples of bulgur prepared commercially the steamed wheat diet. Hutchinson et al. suggest in the United States, and the wheats from which that this increase in food intake could be due to they were prepared, were compared for the effect (I) the destruction of the trypsin inhibitor, (2) of processing on the nutritive value of the wheat. improved palatability, or (3) the treatment having Processing conditions are not reported in detail destroyed the capacity of the wheat to form gluten, but range from continuous pressure cooking to thus making the meal or flour more easily and continuous cooking at atmospheric pressure with efficiently masticated and digested. The last sug­ differing intermediate treatments. The protein gestion was based on the observation that rats (N x 5. 7) contents of the wheat and the bulgur fed barley (which does not form gluten), steamed were not significantly different nor were the PER and unsteamed, showed no significant difference values (method not given). Because of the nature in rate of growth, food intake and efficiency of of the survey, statistical analysis was not possible. utilization. In summary, Pence et al. ( 1965) consider that, Rhi::opus oligosporus fermentation Tempeh in general, the nutritional values quoted in the is an Indonesian product prepared by fermenting literature for wheat may be used for bulgur. soya beans with species of the genus Rhi::opus. Shepherd et al. (1965) compared bulgur Wang et al. ( 1968) compared the PER of: (A) produced from four different wheats: (A) by a casein; (B) wheat, slightly cracked, washed and hot lye treatment followed by a thorough washing, boiled for 12 min, drained and cooled; (C), as (B), (wURLD WHEAT); (B) by a conventional method, followed by inoculation with a spore suspension of with (C) the wheats from which they were made. R. oligosporus NRRL 2710, and incubated at Lye-peeling, which produces a light coloured 31°C for 24 hours; after incubation, steamed for bulgur even from red wheats, did not affect protein 5 min to destroy the mould, freeze-dried and content (N x 5.7). Average PER values were not ground; (D) soybeans, slightly cracked, washed significantly different between wheat and con­ and boiled for 25 min, drained and cooled; (E), ventional bulgur (wheat 1.17; bulgur 1.15) but (0) inoculated and treated as (C); (F), equal the PERs of the lye-peeled bulgur averaged 1.02. weights of (B) plus (D); (G), equal weights of (B) Shepherd et al. (1965) suggest that this reduction plus (D), followed by incubation as for (C). The may be due to the leaching of some of the water­ results are given in Table I 05. soluble proteins and/or to partial destruction of Fermentation of wheat improved the growth lysine. rate and PER significantly but did not have a Steamed Wheat Hutchinson et al. (1964) significant effect on soybean. However, the fer­ compared the growth rate (grams per gram mented mixture of soybeans and wheat produced nitrogen consumed) of male weanling rats, eight a PER comparable to that of casein. per group, over 21 days, on diets containing 1.53% The amino acid composition (by autoanalyzer; Kjeldahl nitrogen, and composed of: (A), un­ tryptophan, microbiologically) and availability heated wheat from a commercial (UK) mill, (in vitro by pepsin-pancreatin digestion) was laboratory-milled to about 65% extraction rate; determined on (B), (C), (F) and (G). The essential 134 MONOGRAPH IDRC-02le

TABLE 105. Rat PERs of wheat and soybean fermented bread fortified with DL-lysine HCI was 11 % by R. oligosporus (data from Wang et al. 1968). (range 2.4 to 21.8%) and with L-lysine HCI the loss averaged 32% (range 25 to 36.8%). The lysine PER content fell between 5 and I 0% during toasting. Rosenberg and Rohdenburg (1952) demon­ (A) Casein 2.81 ±0.10 strated a significant increase in PER by the addi­ (B) Wheat, boiled .28±0.05 J tion of 0.3% L-lysine. (C) Wheat, fermented I. 71 ±0.05 Sure (1954) in rat feeding tests reported the (D) Soybeans, boiled 2.17±0.03 (E) Soybeans, fermented 2.27±0.05 nutritive values of whole rye flour to be markedly (F) Soybeans+ wheat (I: I), boiled 2.49±0.04 superior to whole wheat flour when fed at 9, 8 (G) Soybeans+ wheat (I: I), fermented 2. 79±0.04 and 5% protein levels over a period of I 0 weeks. Additions of L-lysine and L-lysine plus DL­ threonine improved the growth response to whole amino acid composition of wheat and wheat plus wheat flour at an 8% protein level. Little growth soybean was not significantly altered by fermenta­ response was noted when lysine and threonine tion but the available lysine in wheat was increased. were added to whole rye flour at the 8% protein Wheat fermented for 12, 24, 48 and 72 hours level. was also incorporated into rat feeding diets. On Sabiston and Kennedy ( 1957) compared whole the basis of PER value and growth of the rats, the wheat bread and its unbaked ingredients con­ optimal fermentation time for wheat by Rhizopus taining additions of 0, 3, 6 and 12% nonfat dry was 48 hours. milk and 0.166% L-lysine, with white bread con­ Enzyme Treatment Although wheat bran taining 6% nonfat dry milk. Rat growth studies contains about 17% protein, on a dry basis, indicated that white bread containing nonfat monogastric animals are able to utilize only about dry milk was superior to whole wheat bread with­ 60 to 70% of the protein (Saunders et al. 1972). out milk and somewhat inferior to whole wheat Different cellulolytic enzymes available com­ bread containing 3% nonfat dry milk. The PER mercially in the USA were used to process wheat of whole wheat bread containing 0.166% L-lysine bran and one of these (Pectinol 41 P) increased (roughly equivalent to the amount of lysine in 6% the nutritive value of bran as determined by rat nonfat dry milk) was similar to that of bread con­ feeding. taining 6% nonfat dry milk. The PERs of all Supplementation with baked bread were approximately 20% lower than the PERs of their unbaked ingredients. Other Protein Sources Stromnaes and Kennedy ( 1957) reported the Protein supplements and supplementation have PER of bread to be increased by 9%, and of the been considered by several conferences and work­ unbaked ingredients by 11 %. by the addition of ing committees (United Nations 1968; Association up to 6% milk solids. The addition of 0.166% of Food Scientists and Technologists (India) 1969; lysine increased the PER of both the bread and Milner 1969). The text which follows has been unbaked ingredients by roughly 21 %. organized according to the sources of supple­ Jahnke and Schuck ( 1957) reported that the mentation; synthetic amino acids, cereal proteins, addition of three parts of nonfat milk solids and egg and milk proteins, food legume and oilseed 0.25 part of lysine per I 00 parts of flour were protein, protein from microorganisms and fish comparable in protein value to the addition of 12 protein. parts of nonfat milk solids without lysine. Bender( 1957) reported that, when supplemented Synthetic Amino Acids with 0.2% lysine, 1.4% threonine and I. I% Rosenberg and Rohdenburg ( 1951) compared methionine, British bread gave an NPU value of American bread containing shortening, sugar and 80. The NPU values were 46 and 57 respectively dry skim milk with similar breads fortified with for bread alone and bread to which 0.2% lysine 0.5% DL-lysine HCI and 0.25% L-lysine HCI. The had been added. loss of lysine in the standard during baking was Hutchinson et al. (1958) demonstrated that found by microbiological assay to average 15% lysine is the first limiting and threonine the varying from 9.5 to 23.8%. The average loss in second limiting amino acid in unsupplemented HULSE/LAING: PROTEINS OF TRITICALE 135 wheat bread. Rat body weight gain on unsupple­ amino acids had been added. Added glycine and mented bread was about 20% of that on an egg lysine produced a decrease in gas production protein diet. When lysine and threonine were which, Zentner suggests, resulted from the forma­ added to bread to the point that they were no tion of N-substituted glycosylamines by amino longer limiting, growth efficiency rose to approxi­ acids-reducing sugar interaction. mately 85% of the egg protein diet. Yang et al. (1961) studied first the effect in rats Culik and Rosenberg (1958) compared white of graded additions of lysine to wheat flour and bread containing 6% nonfat dry milk with bread second, whether a single daily dose of lysine supplemented with 0.25% L-lysine HCI. The administered by stomach tube was nutritionally comparison was carried out through five succes­ equivalent to the same amount of lysine included sive rat generations. Lysine supplementation in the diet. improved reproduction and lactation significantly Between 0.2 and 0.25% lysine improved body and uniformly throughout the seven litters of the weight gain though certain adverse effects were parent generation and through all successive noted at the I% lysine addition. No difference in generations. Breeding performance on the com­ growth was noted between lysine in the diet and mercial bread diet was inferior to the lysine lysine fed by stomach tube 4, 8, 12 or 16 hours supplemented diet. Bread supplemented with after normal feeding. 0.25 L-lysine HCI as the sole source of dietary Barness et al. (1961) compared white wheat protein maintained near normal reproduction and flour (commercial "cream of wheat") as the chief lactation. source of protein in the diets of 22 malnourished Brown et al. (1959) compared a commercial male Latin American infants aged 3 to 17 months white bread and three types of commercial high in a children's hospital in Texas, in diets (A) with protein bread in rat growth tests in which bread and without milk, (B) with and without lysine provided the sole source of protein. Bread was supplementation. Following an adjustment period, tested with and without lysine supplementation, nitrogen balance studies were carried out for nine the percentage of protein from wheat varying days. from 89 to 60% and from milk and soy proteins Lysine in the "cream of wheat" was determined from 11 % to 40%. The PERs were found to be a as 135 mg/g N by an unstated method. L-lysine direct function of the lysine content of the HCI was added to yield a total of 0.55% of the mixed protein regardless of whether the lysine was wheat. The diets also included starch and sugar, added as L-lysine HCI or as a constituent of milk and the infants received a multivitamin mixture or soya. PER values above 2.0 were recorded daily. Caloric intake varied from 75 to JOO to 120 only when L-lysine exceeded 200 mg per gram of kcalories/kg per day and protein (N x 6.25) from nitrogen. 1.2 g to 4.0 g protein/kg per day. In the wheat­ Hutchinson et al. (1960) also confirmed that, milk mixture 70% of the protein was obtained when fed white bread as the sole source of protein, from the wheat and 30% from the milk on which the mean rate of growth of weanling rats was diet the infants remained in N equilibrium even directly proportional to the mean intake of lysine. when total protein was less than 2 g/kg per day From the results of the experiment, in which the and caloric intake 75 kcal/kg per day. Supple­ protein content of the diet was between 12 and mentation with lysine and potassium gave no 13% and the amounts of added lysine ranged improvement. from 0 to 0.25%, Hutchinson et al. related the A positive effect of supplementation was con­ rate of growth y (grams per rat per day) to the sidered to exist if the N balance during the intake of lysine x (milligrams per rat per day) by supplemented period differed by more than I 0% the empirical equation y = 0.06154x + 0.03. Sub­ of the N intake from the most positive control stitution in the equation of data from growth rates period; a negative effect if the control period comparing bread crumb with bread crust indi­ exceeded the supplemented period by more than cated a loss of 20% lysine in the crust. The slightly I 0% of the intake, and no effect if the balances lower rates of growth recorded suggest a reduction were between those limits. in available lysine during processing of wheat germ. Of the two infants fed wheat alone at 75 Zentner (1961) reported an increased browning kcal/kg per day and protein at approximately of bread crust baked from wheat flour to which 2.0 g/kg per day, one showed increased N reten- 136 MONOGRAPH IDRC-021e tion and one no effect when lysine was added. ration of 150 g per child was given in two portions, When the wheat was supplemented with potassium and was provided on 150 school days in a total alone, there was an increased retention in all three period of261 days. Some 450 children were divided periods studied, and when both lysine and potas­ into four age groups for each sex and examined sium were added, there was increased retention in for height and weight before the feeding program two of the three periods, and no effect in one. In began, at a mid point, and at the end. King et al. the baby who showed increased N retention from reported that although the increments in stature, lysine alone, the effect was much greater with the weight and mean corpuscular haemoglobin con­ double addition of lysine and potassium. centration "are not impressive", they were, in With five babies fed wheat alone at I 00 kcal/ certain cases, statistically significant, indicating kg per day and protein at 2.0 g/kg per day, there some benefit from lysine supplementation. Marked was no effect in two periods when lysine alone was weight gains occurred even in the children fed added, no effect in the three periods when potas­ bread without lysine, compared to those in the sium alone was added, but a significant effect when control school. both lysine and potassium were added. Hutchinson et al. (1963) reported when dried When wheat alone was fed to four babies at bread crumb provided 85% of the diet and the sole 100 kcal/kg per day and protein levels of source of protein for rats that the addition of 0.5% respectively, 2.5, 3.5, 4.0, and4.0g/kgperday, only L-lysine and 0.2% of L-threonine improved bread the baby receiving the 2.5 g protein level showed a protein quality to the rank of "first class". The positive effect in N retention with potassium supplemented bread crumb contained a total of supplementation and an even more marked effect between 0.75 and 0.80% lysine and between 0.5 after the addition of lysine and potassium. and 0.6% threonine. Equivalent levels of lysine Potassium balances determined in 14 babies and threonine required additions to the bread indicated a relationship between increased potas­ crumb of 6.2% casein, 8% egg albumin, 21 % soy sium retention with increased N retention. flour or 18% skim milk powder. It should be noted Positive nitrogen balances were obtained when that whereas the addition of lysine and threonine wheat was fed alone at total protein levels of 2 to did not affect the protein content (12.5%) of the 4 g/kg per day. The data indicated that wheat dried bread crumb, the additions of casein, egg supplemented with lysine and potassium, as albumin, soy flour and skim milk raised the determined in the present study, is an adequate protein to 18.6%, 20%, 18.6% and 17.2% respec­ source of protein for growing infants for the period tively. covered in the study. The lysine and potassium Gates and Kennedy (1964) fed to rats unbaked were effective only when mixed with the daily bread ingredients containing 3, 6 and 12 parts of wheat diet. nonfat dry milk per JOO parts of flour with and Pomeranz (1962) did not find any significant without 0.25 part of L-lysine HCI, and the cor­ correlation between the lysine and protein contents responding breads. The unbaked ingredients with­ of flours of equivalent extraction rates, but higher out added lysine showed significantly increasing extraction flours contained significantly higher PER values with increasing milk solids. The lysine contents. Small losses of lysine in the crumb addition of lysine resulted in significant increases occurred in bread made from commercially milled in PER both with unbaked ingredients and with flours enriched with soy flour, yeast, gluten or bread at all levels of added milk, the proportional wheat germ. Pomeranz suggests that protein­ increase declining as the level of milk solids in­ fortified breads should be classified by their creased. At all levels of milk, and in samples with protein and lysine contents rather than by a and without lysine, increases in PER were description of the level of fortification. significantly inferior in animals fed bread as com­ King et al. (1963) studied the effect of feeding pared with the unbaked ingredients, the average bread, made from white wheat flour enriched to decrease in PER when the ingredients were baked United States standards and the same bread into bread being of the order of 21 %. additionally fortified with L-lysine HCI, at the level Jansen et al. (I 964a, b) demonstrated that PERs of 625 mg/ I 00 g flour to children aged 6 to 18 years of both unsupplemented bread and bread supple­ in two villages in Haiti. The bread contained 4.0 mented with 0.3 of L-lysine HCI per JOO g offlour to 4.6 mg L-lysine HCI, which was consistent with were reduced as baking time increased from 0 to an expected loss of 15% during baking. The daily 50 min. Of the added lysine, 30% became un- HULSE/LAING: PROTEINS OF TRITICALE 137

TABLE 106. Protein content of macaroni, total L-lysine HCl in protein, mean protein intake and PER of plain and enriched macaroni, with and without added lysine HCl, on rat PER (data from Bains et al. 1964).

Total Protein L-lysine Mean content of HCl in protein macaroni protein intake (%) (%) (g) PER

(A) Plain macaroni 12.0 3 .16 19.59 l.10 (B) (A) diet plus L-lysine HCl 0.25 g/100 g 12.0 5.40 26.95 2.1 l (C) Enriched macaroni (plus casein and skim milk solids) 17.0 6.80 28.98 2.48 (D) (C) diet plus L-lysine HCl 0.25 g/ l 00 g protein 17.0 9.60 29.08 2.63 available after baking for 30 min at 450°F flour and mixtures with and without added lysine (232°C). When the baking time was reduced to and other amino acids. 20 min no significant nutritional loss was found by Yamazaki (1968) recommended that bread sold rat growth assay though an 18% loss was indicated in Asia should be fortified by lysine up to 0.25% by ion exchange chromatography. In bread with­ of the flour weight. out milk powder 15% of the added lysine became Maleki and Djazayeri ( 1968) studied the supple­ unavailable when the bread was baked for 30 min mentation of Arabic bread with lysine, threonine at 450°F (232°C). The loss of lysine during and methionine. They found the PER (0.57) of baking increased significantly in the presence of non-supplemented flour to decrease to 0.4 during nonfat dry milk. When additions of between 6 baking. The addition of 0.3% L-lysine raised the and 14% of nonfat dry milk were made to doughs PER of flour to 1.4 and of bread to 1.5, and 0.6% baked for 20 min at 450°F (232°C) the loss of threonine added before baking raised the bread lysine was about 36%. PER to 2.3. Methionine produced no significant Bains et al. (1964) compared rat PERs of: (A) change. macaroni made from Indian durum wheat semo­ Hedayat et al. (1968) studied the biological lina, (B) diet (A) fortified with 0.25 g L-lysine HCl value of Iranian commercially milled whole wheat per 100 g protein after processing, (C) enriched flour, with and without additions of L-lysine HCl macaroni made from durum wheat semolina, when baked into Iranian village type bread. The casein, and skim milk solids in the weight ratios flour was sifted by the baker before use to remove of 90: 7: 3, (D) diet (C) fortified with 0.25 g about 5% of the bran and coarser particles. L-lysine HCl per 100 g protein, after processing. The NPU of unenriched whole wheat flour was The macaroni was included in 10% protein rat found to be 42.0 and of village bread 42.5. The diets fed ad libitum over 28 days. The results are equivalent Biological Values were 46.0 and 46.5. given in Table 106. The NPU of the flour with 0.2 g L-lysine HCl per Addition of lysine to plain macaroni in diet (B) 100 g flour was 50.5 and of the village bread 60.0. significantly improved the quality of the protein The equivalent BY were 55.5 and 67.0. but fortification with milk protein and casein in Food consumption and growth rate were higher diet (C) effected a still greater improvement. The in the rats fed bread from enriched flour than in PER of macaroni in diet (C) was not significantly those fed bread from unenriched flour. improved by the addition of lysine. Rats fed diet Hedayat et al. also found that weanling male (B) had significantly lower percentage of fat in Wistar rats showed significantly higher serum their livers than those on diet (A). albumin, liver fat and liver nitrogen when fed Howe et al. (l 965a, b) discuss the effect on PER whole wheat flour supplemented with 0.2 g L­ of supplementing a wide range of cereals with lysine HCl/l 00 g flour than those fed unsupple­ protein sources including fish flour, peanut flour, mented wheat flour. No similar differences were sunflower, cottonseed flour, sesame flour, soybean found among female rats. 138 MONOGRAPH IDRC-021e

The effect of the supplementation with L-lysine as a preliminary period, faeces and urine being and DL-threonine, of Indian wheat and of a poor collected in the second five days. The order in wheat diet, was studied in groups of eight 20-day­ which the diets were fed was (I) basal wheat, old male albino Wistar rats by Daniel et al. (2) wheat plus 1.5 g L-lysine HCl/child per day, (I 968b). The lysine and threonine contents of the (3) wheat plus 0.86 g DL-threonine/child per day, wheat and of the diets were assayed microbio­ (4) wheat plus 1.5 g L-lysine HCI and 0.86 g DL­ logically. The protein content of the basal wheat threonine/child per day, (5) skim milk powder diet was 10.3% and of the basal poor Indian wheat diet, (6) low protein diet, similar to that of diet (PWD) was 12.3% (N conversion factor Parthasarathy et al. (1963). The proteins of the not stated). The lysine and threonine contents wheat diet contained, in grams/ 16 g N, lysine 3.1 of the wheat were, respectively, 3.0 and and threonine 3.8 (wheat protein containing lysine 2.4 g/16 g N and of the PWD 3.3 and 2.8 g/ 2.4 and threonine 2.8). 16 g N. L-lysine and DL-threonine were added Nitrogen in the diet, urine and faeces was to raise the levels approximately to those in egg determined by micro-Kjeldahl. The digestibility proteins, namely 6.6 g and 4.9 g/16 g N respec­ coefficient and the biological value of the diets tively. PER of the wheat increased from 1.62 to were calculated according to Tasker et al. (1962). 1.99 when L-lysine was added and to 3.02 when The net protein utilization operative (NPU0P) L-lysine and DL-threonine were added. The PER was calculated according to Platt et al. (1961 ). of skim milk powder was 3.13. The PER of the The wheat diet at a level of 1.5 g protein/kg PWD diet when also fortified with vitamins and body weight met all the essential amino acid minerals increased significantly from 2.03 to requirements of children as assessed by Nakagawa 2.58 on the addition of L-lysine and to 2. 70 on the et al. (1962) except for lysine. Nitrogen retention addition of both lysine and threonine. The on the wheat diet was 10.2% of intake. Supple­ difference between the PERs on the addition of mentation with L-lysine raised N retention signifi­ lysine and of lysine plus threonine was not cantly to 21.5% of intake. Supplementation with significant. The PER (I. 73) of the PWD diet when DL-threonine raised retention to 14.4% but this it was not also fortified with vitamins and minerals was not significant. Supplementation with both showed no significant improvement on the PER L-lysine and DL-threonine increased retention to a (1.90) with addition of lysine; the addition of highly significant 30.4% of intake, comparable lysine and threonine improved the PER to 2.18 with 31.3% N retention on the milk diet. which was significantly higher than the basal diet The NPU of the wheat diet was 37.7, which but not significantly higher than the diet supple­ increased significantly to 48. 7 by supplementation mented with lysine alone. It was concluded that with L-lysine and non-significantly to 42.0 by deficiencies in vitamins and minerals must be supplementation with DL-threonine. Supplementa­ corrected in order to gain greatest benefit from tion with L-lysine and DL-threonine significantly supplementary amino acids. increased the NPU to 57.3, comparable to that The effect of supplementing a poor wheat diet from the milk diet of 59. I. with L-lysine and DL-threonine was also studied The protein intake ranged from I. 78 to 1.80 by Daniel et al. (I 968a) in feeding trials with seven g/kg body weight on the different diets. The net boys aged I 0 to 12 years living in a boarding house available protein in grams per kilograms body in Mysore, India. The basal wheat diet was that weight was: wheat diet 0.67, wheat plus L-lysine customarily eaten in the home and provided daily 0.80, wheat plus DL-threonine 0.75, wheat plus 44. 7 g protein (N x 6.25), of which whole wheat L-lysine and DL-threonine 1.04, milk diet 1.11. flour contributed 33.3 g, red gram dhal (Cajanus These were compared with the Food and Agri­ cajan) 3.4 g, skim milk powder 1.8 g, and vege­ culture Organization (1965) reference protein tables and condiments 6.2 g. A diet based on skim requirements of0.72 g/kg body weight. milk powder in which maize starch and skim milk The effect of supplementing wheat with L-lysine powder were substituted for the whole wheat flour HCI in the all-vegetable diet of children aged 2 to and the red gram dahl in the wheat diet was used 5 years in an orphanage in Vellore, South India, for comparison. was studied over six months by Pereira et al. ( 1969), The experiment comprised six periods each of the children being divided into two matched 10 days duration, the first five days being treated groups, (A) experimental with lysine supplementa- HULSE;LAING: PROTEINS OF TRIT!CALE 139 tion and (B) control, without supplementation. report that 25% of }he added lysine was lost in the The basic orphanage diet provided 2 g vegetable bread but only 4% in the chapati. protein and 100 kcal/kg body weight per day. The Jansen ( 1969) reported adding several protein wheat was fed as wheat flour and broken wheat supplements to bread. The protein content (dry grains, boiled, roasted or fried, and provided 54/0 weight basis) of the unsupplemented bread was of the daily calories and 85% of the daily protein. 13% and the net protein value (NPV) 7.3. The The only difference in the diets of the two groups addition of 4% nonfat dry milk raised the protein lay in the lysine supplementation of the wheat content to 16% and the NPV to 8.5; 4.1% toasted portion of the experimental group's diet. From soya increased the protein to 17% and the NPV analyses (by autoanalyzer) of samples of cooked to 9.0, and 10% nonfat dry milk raised the protein foods, children in the control group had an intake to 17% and the NPV to 10. The addition of 0.3% of 0.54 g lysine per day, those in the experimental L-lysine HCl did not affect the protein content group 0. 73 g lysine per day. The losses of lysine but raised the NPV to 10.0, the equivalent of adding during cooking were 21 % in the control group 10% nonfat dry milk. and 30% in the experimental group. Efremov et al. ( 1970) described how five middle­ At the end of the six months, the children in the aged men employed in the baking industry of the lysine-supplemented group had grown significantly USSR were fed a standardized mixed diet of 3400 taller than those in the control group. Statistically Calories per day containing 500 g bread over significant differences in weight were not observed, five successive 10-day periods. Only the composi­ both groups losing weight in the last month of the tion of the bread in the standard diet was changed trial which occurred during the hottest part of the for each 10-day period. During the first 10 days year. No statistically significant differences were they received ordinary wheat bread (composition observed between the groups in haemoglobin, not recorded); during the subsequent periods the packed cell volume, total serum proteins, serum bread contained, per 100 g flour (A) 10 g dried albumin, retention of absorbed nitrogen, or daily skim milk, (B) 5 g fat-free fish flour, (C) 20 g creatinine excretion. Nor were significant dif­ soybean (composition not stated), (D) 0.5 g lysine ferences observed in minor illnesses or general HCI. The nature and composition of the non­ health. bread portion of the diet is not given but the Mitsuda ( 1969) described how bread fortified tabulated data indicates the bread provided about with lysine was tested among groups of Japanese 40% of the total nitrogen intake. The subjects' school children. The bread was enriched with 0.8 g body weight did not change and nitrogen balance of L-lysine HCl per school lunch serving to offset remained positive throughout. The methodology the loss of lysine during baking. At this lysine is not described but Efremov et al. quote the level excessive browning of the bread together Indices of Net Utilization of Protein (%) as with a slightly bitter taste was noted. The loss of follows: bread (control) 12.6, (A) plus skim milk lysine was reduced by baking at a lower tempera­ 10.0, (B) plus fish flour 15.7, (C) plus soya 17.3, ture for a longer time. (D) plus lysine 15.1. Adrian and Frangne ( 1969) studied biscuits The quoted skim milk result appears to conflict baked from wheat flour plus gluten enriched with the conclusion that the addition of skim milk, alternatively with groundnut (peanut) meal, fish fish flour, soya or lysine to bread improved the meal, skim milk powder or lysine. During baking efficiency of protein utilization. the PER of groundnut- and fish meal-enriched Clark et al. ( 1970) determined the requirements biscuits fell by 30% and 40% respectively. Greatest of adult human subjects 20 to 25 years old for losses occurred in the milk-enriched biscuits. methionine and cystine using white wheat flour Adrian and Frangne state the loss of amino acids as the dietary base. There were six men in the from the "digestible" fraction to be less than is first experiment and five young men and two accounted for by the drop in PER. It would women in the second experiment. Amino acids appear that lysine was rendered unavailable by were determined by autoanalyzer. interaction, possibly with reducing sugars. Part of the amino acids were provided by the Matthews et al. ( 1969) compared the effect of flour and the remainder in crystalline form. Total adding lysine to chapati and bread made from daily nitrogen intake was 6.0 g per person. The wheat flours of 93% and 85% extraction. They requirements varied from 700 mg methionine plus 140 MONOGRAPH IDRC-02le

400 mg cystine to 260 mg methionine plus 280 control groups, but the lysine groups showed mg cystine. The sulphur-containing amino acids significantly higher gains in mid-arm circum­ present in 142 g white wheat flour appeared to be ference than the no-lysine group. adequate for five of the six men in the first experi­ The point is made that the final measurement of ment. weight and/or arm circumference gain was taken Hedayat (1971) and Hedayat et al. (1971) in mid-spring when food was most scarce in the describe a school lunch study conducted among villages; this could explain the considerable children aged from six to 12 years of both sexes number of children in the control group who in five Iranian villages over eight months in the showed loss of weight. school year 1968-69. Children in three villages Hedayat et al. ( 1973) discussed the possible received bread enriched with lysine, vitamins and causes of the essentially negative results of the minerals. In the fourth village the lunch included enrichment of bread with lysine and concluded that bread plus vitamins and minerals but without since the provision of school meals had a very added lysine. In the fifth village, neither lunch nor positive result in improving weight and anthro­ bread was provided. The bread was made locally pometric measurements in children compared to from Iranian flour of 95% extraction enriched those who did not receive school meals, school with a vitamin-mineral mixture alone, or with the feeding programs might be more beneficial than same mixture and lysine HCI to give 2.3 g of lysine lysine enrichment. per kilogram of flour. Chemical and biological Ghai and Chaudhuri (1971) studied the effect of tests on rats conducted earlier (Hedayat et al. supplementing wheat flour with lysine in the diet of 1968) had shown that most of the added lysine malnourished preschool children in India. The 44 was still available after baking into bread. children, aged one to six years, though free from The bread was served at breakfast ( 125 to 150 chronic and acute diseases, had weights below g) and with lunch ( 150 to 200 g). The lunch meals 60% of the mean weight for age standards. They provided approximately 50% of the recommended were divided into two groups of 22, matched for dietary allowances (National Academy of Sciences age, sex and weight. The diet provided 120 (US) 1964) for protein, calories, calcium and Calories and 2 g protein per kilogram body weight, ascorbic acid, 66% of the thiamine, 85% of the all the protein and most of the Calories being riboflavin and niacin and 199% of the iron. The provided by the wheat flour. The allowances were composition of the lunch dishes varied according adjusted once a week to allow for weight gains, to the seasonal availability of vegetables. Animal and the observations were carried on for a period protein and pulses were excluded, the intention of two months. being to ensure a limitation in lysine. There was no One group received the diet without supple­ control of food eaten by the children in their homes mentation, the other wheat supplemented with but a food consumption survey indicated that 0.1 % lysine. The level of 0.1 % was chosen because dietary protein was limiting in lysine in 97% of studies at the Nutrition Research Laboratory, the relevant village households surveyed. In total, Hyderabad (1967-68) had indicated that lysine 203 children received lunch and lysine-supple­ supplementation at the 0.1 % level increased the mented bread, 90 children received lunch with PER of wheat flour from I . 78 to 2.15 but that no bread without lysine supplement and 117 children further increase occurred when the level of lysine received no lunch or bread. The children received a was raised to 0.2%. standardized clinical examination, and height, The differences found between the two groups weight, skinfold thickness of the triceps and arm in the rate of weight gain, changes in serum circumference were measured. proteins, and ratio between non-essential and The children receiving the school meal showed a essential amino acids in the serum were not marked gain in weight and mid-arm circumference significant. compared with the control group, but no significant Oiso ( 1971) reviewing clinical studies of amino difference in weight gain between the lysine and acid fortification in Japan refers briefly to studies no-lysine groups was noted. The group receiving on adult males on a regimen consisting of 600 the school meal without lysine showed significantly g bread and 60 g sucrose. Whether receiving higher height gains compared with the lysine and supplements or not, the men remained in negative HULSE/LAING: PROTEINS OF TRITICALE 141

N balance though supplementation with L-lysine an equivalent amount of nitrogen from glycine. HCI tended to improve the N balance and Amino acids were added to give 270 mg lysine, diminish fatigue. 90 mg tryptophan, 270 mg methionine, 270 mg Young and Scrimshaw (1971) reviewed clinical isoleucine, 270 mg valine and 180 mg threonine/g studies in the United States on the amino acid N in the diet. The results suggest that the main fortification of protein foods. They cite Hegsted improvement in protein quality resulted from the et al. ( 1955) who found a positive effect of methio­ addition of lysine although the addition of trypto­ nine supplementation at 1.0 g/day in young phan and methionine further improved nitrogen women fed an all-vegetable protein diet in which retention. wheat provided 48% of the total nitrogen, and rice Bressani ( 1971) considers that some of the and other cereal products the remainder. Hegsted effects observed with the mixture of amino acids et al. (1955) also found that 0.9 g/day supple­ may result from their low biological availability in mentary lysine improved nitrogen retention. The wheat or to certain imbalances among them. unsupplemented diet provided about 1200 mg Supplementation may not be beneficial if the lysine/day. amounts of individual amino acids added are Young and Scrimshaw ( 1971) question whether greater than the optimum, thus giving rise to an the positive effect of supplementary methionine overall imbalance. would have continued during a longer balance To determine the optimum level of lysine to be period since the intake of total sulphur-amino added to wheat flour, balance tests were carried acids provided by the unsupplemented diet ap­ out at constant caloric intake on three children at peared adequate. They cite Hoffmann and McNeil two levels of dietary protein. At a dietary protein ( 1949), who found that the nutritive value of wheat intake of 2.0 g/kg per day, 61 mg lysine per gram gluten fed to adult men could be significantly nitrogen was sufficient to increase nitrogen reten­ enhanced by supplementation with L-lysine. Using tion. At a protein intake of 3.0 g/kg per day, 107 the nitrogen balance index (NB!) method de­ mg lysine per gram nitrogen were needed. The veloped by Allison (1959, 1964), they found, with ratio of lysine need to nitrogen intake was nine subjects, that 4% L-lysine added to the protein similar for both levels. improved the NB! from 0.62 ± 0.09 without Reddy (1971) conducted nitrogen balance supplementary lysine, to 0.76 ± 0.08, with supple­ studies among Indian children, from two to five mentary lysine. The latter figure compares with a years, who were moderately undernourished value of about 0.73 for casein (Allison 1959). though not showing signs of severe protein Bressani (1971) summarized earlier work malnutrition. The diet, composed of wheat flour (Bressani et al. 1960; Bressani et al. 1963) on the of protein content 10.7% (conversion factor not amino acid supplementation of Guatemalan wheat stated), safflower oil and sugar, was fed to provide flour, of 70% extraction, in nitrogen balance 2 g protein and I 00 kcal/kg body weight. Vitamin studies in children. The essential amino acid and mineral supplements were also provided. composition of the flour is given as, in milligrams Baking the flour into chapatis did not give rise per grams nitrogen: arginine 229, histidine 155, to a significant loss in available lysine (method of isoleucine 239, leucine 375, lysine 159, methionine Carpenter, 1960). The lysine content was assayed I0 I, cystine 112, phenylalanine 324, tryosine 191, micro biologically (Steele et al. 1949), from a water threonine 183, tryptophan 52, valine 245. The basal extract of the fortified wheat; the free lysine in the diet in percent by weight consisted of: wheat 85, water extract was considered to be available. The gluten 7, glycine 3, maize starch 5, to give a experiment was divided into three I 0-day diet protein content (N conversion factor not stated) of periods: Period I, an unfortified wheat diet;

18.7 and 385 calories/I 00 g. In most of the studies Period II, wheat diet supplemented with 0.1 / 0 the children received each diet for a total of nine lysine HCI; Period III, unfortified wheat diet. days with a 4-day adjustment interval between The unsupplemented wheat diet provided 56 mg treatments. Three balances were obtained per lysine/kg body weight; the supplemented diet 72 dietary treatment. mg lysine/kg body weight. All of the children The weight of amino acids replaced an equal gained weight over a period of four weeks and in weight of maize starch and the nitrogen replaced all the nitrogen retained during the three periods 142 MONOGRAPH IDRC-02Ie was essentially similar. The suggested protein ( 1957). The nitrogen balance was the principal requirement for infants is about I. 7 g/kg and for method of evaluation. pre-school children about I g/kg (World Health The mean nitrogen balances achieved on the Organization I 965a). Snyderman et al. ( 1959) wheat diet were (I) with no supplementation, reported the lysine requirements for infants to be -2.38gN; (2) with 4.0gNSN, -0.81 gN; (3) about 90 mg/kg body weight. Assuming that with 8.0 g NSN, -0.12 g N. These results confirm amino acid requirements run parallel to protein earlier findings that NSN supplementation has a requirements, the calculated lysine requirement for sparing effect on the protein requirements of pre-school children would be 53 mg/kg body humans. weight which is close to the 56 mg/kg in the un­ The importance of glutamic acid as a source of supplemented wheat diet. This may explain the non-specific nitrogen in diets for rats on an amino absence of any beneficial effect from lysine supple­ acid diet was tested by Hepburn et al. (I 960a). mentation in the children discussed above. Male weanling rats when fed a diet containing an Young and Scrimshaw ( 1971) reviewed clinical amino acid mixture patterned on that of wheat studies in the United States on the level and kind gluten, and supplemented with histidine, lysine, of non-specific nitrogen in the human diet and methionine, threonine and tryptophan showed an emphasized the need for extensive additional unusually high rate of weight gain. When the non­ clinical studies into the contribution which the essential amino acids (except cystine and tyrosine) "non-essential" amino acids may take, in extend­ were eliminated from the diet, it was found that the ing available protein supplies. omission of glutamic acid in each instance resulted Kofranyi ( 1971) following long-term nitrogen in a reduced growth rate. The elimination of other balance tests on human subjects, also states that non-essential amino acids did not produce the the biological value of proteins is not determined same effect. When increments of glutamic acid by the limiting essential amino acids alone but also were added to glutamic acid-free diets, progres­ by the proportions of the amino acid mixtures sively greater weight gains were obtained. Maxi­ including sources of "non-essential" nitrogen. mum growth rate was obtained at the 5.66% level Kies et al. ( 1972) reported on nitrogen balance of glutamic acid. Food intake was also decreased studies in 10 adult men fed diets providing equal, at low levels of glutamic acid, perhaps as a result but sub-optimal, amounts of protein (4 g nitrogen of reduced palatability in the absence of glutamic per subject per day) from rice, maize, wheat or acid, or loss of appetite due to amino acid im­ milk plus, in each case, 0, 4.0 or 8.0 g N/day from balance. non-specific nitrogen (NSN) consisting of an Taylor et al. ( 1972) examined the effect of lysine isonitrogenous mixture of glycine and diam­ supplementation of wheat gluten in young male monium citrate. The wheat source was enriched students at the Massachusetts Institute of Tech­ wheat flour with a nitrogen content of 0.0184 g N nology. Two levels of protein intake, 0.27 g and per gram flour, fed as drop biscuits. The experi­ 0. 73 g protein/kg per day and two levels of caloric mental plan consisted of three successive 27-day intake were studied in seven subjects and eight periods. Each period consisted of an introductory subjects respectively. Caloric intake was either 2-day nitrogen depletion phase (2.0 g N per sub­ adequate for weight maintenance or 80% of the ject per day), a 5-day nitrogen adjustment phase adequate level. Mean nitrogen balance at low (4.0 g N per subject per day) and four experimental protein and adequate caloric intake was, in phases of5 days each. These are considered to be of grams nitrogen per day, - 1.29 without, and sufficient length to give data for accurate com­ - 1.05 with, lysine supplementation. At low parison (Kies et al. 1967; Kies and Fox I 970a). protein and inadequate caloric intake the cor­ The sequence of the three periods and the order responding balances were - 2.02 and - I. 70 g N of the experimental phases within each section per day. At the higher protein intake, lysine were randomized for each subject. Caloric intake improved nitrogen balance by +0.43 g at both for each subject was maintained at the level levels of caloric intake. required for approximate weight maintenance by Tara et al. ( 1972) used atta (milled cleaned com­ varying maize starch and fat intake. The essential mercial wheat) to prepare chapatis and baked amino acid content provided by 4.0 g N from the yeast-raised bread, without and with additions of wheat was estimated from tables by Orr and Watt L-lysine HCI. They found a loss of lysine of 12.5%, HULSE/LAING: PROTEINS OF TRITICALE 143

as assayed microbiologically, in yeast-raised loaves quality of proteins. This is especially true of fortified with 0.2% lysine HCI and a loss of 5.5% cereal and vegetable diets ... " in the unfortified loaf. Loss of lysine m the chapatis was negligible. Cereal Proteins Biological testing was carried out using 21-day Bakers of bread have mixed other cereals with old weanling male albino Wistar rats in groups of their wheat flours for as long as their art is 10, over a period of 4 weeks. Each diet contained recorded. The ancient Egyptians and Romans amounts of baked or unbaked wheat ingredients mixed barley, oats or rye with wheat flour and to provide about 10% protein (N x 5.7) and the pioneers in North America mixed upwards of25% animals were fed ad libitum. rolled oats with wheat flour to make flat breads Differences between the PER of the chapatis which they baked on griddles over open wood and the unbaked ingredients were statistically non­ fires. significant. The PER of the chapatis fortified with The protein content of wheat flour can be 0.15% L-lysine HCI was significantly higher (38%) concentrated in at least two general ways (I) than that of the unfortified chapatis. concentration of the endosperm protein by re­ The microbiological finding that the loss of moving part of the predominant starch; (2) con­ lysine on cooking chapatis was negligible was centration with selected high protein fractions confirmed by the finding that the PER of chapatis from the bran and heat-stabilized germ. fortified with lysine before cooking (2.04) and the Wheat Gluten. Bran and Germ Wheat endo- PER of those fortified after cooking (2.13) were sperm protein can be concentrated by either wet little different. The animals fed lysine-fortified or dry processes. The "wet" processes depend diets ate more food than did those on unfortified upon washing out the starch from flour-water diets. doughs leaving a concentrate of gluten which can Although a loss of 12.5% in lysine content on be added subsequently to bread doughs either in baking bread was found microbiologically, and the wet state or after drying. The "dry" concentra­ the PER of bread fortified with 0.2% lysine HCI tion processes depend upon controlled grinding before baking was 2.21 compared with a PER of and fractionation of the wheat components. 2.34 for bread fortified after baking, this difference Research on wheat proteins began in the in PER was not statistically significant. The PER Institute of Anatomy and Chemistry at Bologna, of the fortified bread was however about 60% Italy, where Beccari (1745), the professor of higher than that of the unfortified bread and the medicine, described how he separated gluten from amount of food consumed by the rats on the a wheat flour dough by washing out the starch lysine-fortified diets was higher than that eaten by with water. Patents for the production and utiliza­ the rats on the unfortified diets. tion of wheat gluten in bread, biscuits and other Hegsted ( 1971) stated there is abundant evidence foods were reported in the middle of the 19th that when an amino acid is added to a diet con­ Century, e.g. Johnson (1853). taining a nutritionally inadequate supply of that Supplementation of wheat protein with wet or amino acid, the rate of weight of young rats is dry gluten increases the total proportion of protein improved. He considers that none of the assay but does not greatly alter the balance of amino techniques so far available are satisfactory for acids. The protein content of dried washed crude evaluating the relative potency of protein for gluten ranges from 75 to 85% (Kasarda et al. maintenance. He is doubtful that assays obtained 1971). Hale (1963) describes how to make high with young rats are valid estimates of the utility protein bread from added wheat germ and gluten of proteins for man, either adult or growing child, flour of 41 % protein content. Fance and Wragg because during all stages of postnatal development (1968) describe how "gluten bread" containing a much larger proportion of dietary protein is not less than 16% protein and "high protein utilized for maintenance than for growth. Hegsted bread" containing not less than 22% protein on a has said: "A minimum conclusion, it seems, would dry weight basis can be made to British statutory be that amino acid fortification programs based standards. upon rat assay alone would be foolish if not Results which indicate that the proteins of dangerous". " ... amino acid scores appear to be wheat bran and germ are biologically superior to quite inadequate estimates of the nutritional endosperm proteins were summarized by Mitchell 144 MONOGRAPH IDRC-02le

TABLE 107. Effect of adding wheat germ and gluten on protein and lysine content of bread baked from three flours (data from Pomeranz 1962).

Patent Brown Wholemeal

Lysine Lysine Lysine Protein Lysine (%of Protein Lysine (%of Protein Lysine (%of Flour supplement (%) (%) protein) (%) (%) protein) (%) (%) protein)

0 13.8 0.46 3.3 15. I 0.52 3.4 14.6 0.50 3.4 Germ 3% 14.4 0.49 3.4 16.6 0.58 3.5 15.7 0.61 3.9 Gluten 3% 15.2 0.49 3.2 17.2 0.63 3.7 16.7 0.58 3.5 Germ 5% 14.7 0.51 3.5 16.4 0.60 3.7 16.3 0.62 3.8 Gluten 5% 17.0 0.51 3.0 18.3 0.63 3.4 18.7 0.60 3.2 Germ 10% 15.9 0.59 3.7 17. I 0.69 4.0 17.0 0.68 4.0 Gluten 10% 20.2 0.55 2.7 21. 7 0.67 3.1 21.1 0.64 3.0

(1925) and Boas Fixsen and Jackson (1932). By improving protein value was described by Fellers and large, both in developed and developing et al. (1966), Sullivan (I 967a, b) and Rozsa (1968). countries, most of the bran and germ are con­ Bradley (1965) has demonstrated that the milling signed to animal feeds. fraction known as "Shorts" contains roughly Westerman et al. (1952) described the biological double the quantities oflysine and threonine found superiority of defatted wheat germ added at 4 and in an isocaloric quantity of human milk. 6% levels to nonenriched white flour. Rand and Fellers et al. (1966), Sullivan (I 967a, b), and Collins (1958) described the nutritional improve­ Rozsa (1968) described how carefully selected ment of adding IO and 15% of solvent-defatted mill feeds were ground to fine particle size and wheat germ to white flour. subsequently screen-separated to produce wheat Pomeranz (1962) fortified a variety of patent, protein concentrates. Sullivan (I 967a) discussed long extraction (brown) and whole meal flours several such wheat protein concentrates, one of with gluten and wheat germ. The gluten contained which contained 23% protein, approximately 71.7% protein (N x 5.7) and 1.47% lysine, and the 1.05% lysine and 0.74% threonine. Sullivan (I 967b) wheat germ 33. I% protein and 2.24% lysine. The described how a straight grade hard winter wheat results of adding 3%, 5% and I 0% of wheat germ flour was blended with (a) 30% of "Shorts" and and gluten on the bread baked from three flours (b) 30% of a wheat concentrate produced by fine of different extraction rates are quoted in Table grinding and screening "Shorts". These blends 107. were tested in Egyptian flat bread, and in India Pomeranz also reported a high positive correla­ and Pakistan in chapatis, biscuits and other cereal tion (r= +0.91) between the protein and the products. The nutritional values of the blends lysine contents of low extraction flours. were reportedly superior to Indian atta which Howard and Anderson (1965, 1968) described approximates to a whole wheat flour. the treatment of obesity with a commercial Graham et al. (1970) evaluated a mixture of dietetic bread, the "Cambridge Formula Loaf', equal parts of unbleached white flour and a containing 25% protein derived largely from added protein concentrate from wheat shorts, protein stabilized wheat germ. Among I 08 patients, the content 18.8% (N x 6), in feeding trials with mean weight loss was significantly higher over infants and young children. About two-thirds of eight weeks in those fed the high protein, wheat the protein was derived from the shorts, the lysine germ loaf than those fed brown bread or a com­ content of which is higher than the flour. The PER mercial wheat germ bread. of the mixture as obtained was 1.73 ± 0.041 Improved milling technologies offer several (standardized 1.31 c.f. casein 3.27 ± 0.044) and means of concentrating the natural wheat protein the NPU 47, these values being superior to those resource in flour and bread. reported for wheat flour. Wheat Protein Concentrate In the late I 960's All the children received daily vitamin-mineral a new method of concentrating wheat protein and mixtures. The approximate minimum intake of HULSE/LAING: PROTEINS OF TRITICALE 145 milk protein and total calories required by each reported, the proportional loss increasing with child for continued accelerated growth and normal increasing proportion of WPC in the blend. The serum proteins were obtained. Each child received percentage losses in lysine were 2.6, 11.7, 11.9 isonitrogenous and isocaloric levels of the cereal and 18.8% as the WPC was progressively in­ mixture. The test periods of 15 or 30 days were creased from 0 to 45%. Tryptophan fell 2.0% when preceded and followed by 9-day periods in which no WPC was present but thereafter showed no casein was the protein source. loss during baking. As judged by nitrogen absorption and reten­ The rats fed the diets with the unbaked flour tion, rates of weight gain and serum albumin showed a progressive increase in food intake, levels in three infants receiving the wheat flour­ weight gains and liver weights as the proportion wheat concentrate mixture as the sole source of of WPC increased. On average, for each 15% protein, the mixture's protein value was approxi­ replacement of flour with WPC there was an in­ mately 60% that of casein. The mean of 60% N crease of 0.2 in PER. The NPU also increased retention preceding and following casein periods progressively. is close to the 63% found in previous studies with Rye Flour The effect of supplementing rye wheat flour (Graham et al. 1969). Rates of weight flour of98%, 87% and 45% extraction rates with a gain for the same periods gave a mean of 74% range of other foods of animal and vegetable that of casein, higher than the 67% found in the origin was studied by Szkilladziowa ( 1962) in rat wheat flour studies (Graham et al. 1969). In the feeding tests using the methods referred to in her wheat flour studies there was a mean fall of 0.08 previous work ( l 960a, b, 1961 ). A mixture of one and a final level of 4.02 g of albumin/ 100 ml part rye flour and one part wheat flour, both of serum; in the present wheat flour-wheat concen­ 98% extraction gave no improvement over the trate studies there was a mean fall of 0. 77 and a rye flour alone. final level of 3.49 g/l 00 ml serum. When L-lysine was added to the mixture at a level of 48 mg/kg body weight per day in trials Egg and Milk Protein with two infants the mixture's protein value im­ Athenaeus, in his book 'The Deipnosphistai", proved by about 50%. This improvement still left written in 230 AD, tells us that early Egyptian the biological value of the mixture significantly bakers made bread from wheat mixed with cow's below casein whereas with wheat flour alone, milk. Cato also describes how Roman bakers lysine fortification brought the relative biological added milk and cheese to their bread, in addition value very close to that of casein. to nuts and sesame seed. The theoretical improvement in lysine content The high cost of egg proteins restricts their use provided by the wheat flour-wheat concentrate to specialty and highly priced breads and they mixture seems, at least in infants and young make little major contributio:i to bread protein children, to have been offset by inferior digesti­ supplementation. One exception is Jewish "Challa" bility. bread which may contain as much as 2.5% of The effect of substituting a commercial wheat additional protein derived from egg yolk. In protein concentrate (WPC) of protein content many developed countries, skim milk, skim milk 15.35% (N x 5.7) for 0, 15, 30 and 45% of the flour powder (nonfat dry milk), and in some cases in a bread formula on the nutritional value of the buttermilk, are used to supplement bread protein, ingredients and on the baked bread was studied and several countries regulate the quantity to be by Ranhotra et al. ( 1971). Amino acids (except added to such breads (e.g. Government of Canada, tryptophan) were determined by autoanalyzer; Food and Drug Act and Regulations; Government tryptophan by the method ofTkachuk and Irvine of United States of America, Federal Food, Drug (1969). PER and NPU (Miller and Bender 1955) and Cosmetic Act; United Kingdom Govern­ values of a 10% protein diet were determined with ment, Bread and Flour Regulations). six rats per diet, fed ad libitum for 14 days, using Brown et al. ( 1959) and McLaughlan and casein as control. Morrison ( 1960) have reported a highly signifi­ The WPC was lower in protein content, lysine cant correlation between the PER of bread and and fibre, than previous samples examined. A the total amount of lysine present irrespective of loss in all essential amino acids during baking is the source of lysine. 146 MONOGRAPH IDRC-02le

TABLE 108. Effect of additions of nonfat dry milk with and without L-lysine HCI content and PER of bread and the unbaked ingredients (data from Gates and Kennedy 1964).

Nonfat dry milk (parts per JOO parts Nitrogen Adjusted PER by weight of.flour) (%) (Casein= 2.50)

Unbaked ingredients 3 2.67 1.07 6 2.75 1.44 12 2.93 1.87 Bread 3 2.74 0.93 6 2.84 I. 05 12 3.02 I. 35 Unbaked ingredients + 0.25 lysine 3 2.73 2.11 6 2.82 2.11 12 2.98 2.28 Bread+ 0.25 lysine 3 2.76 I. 70 6 2.91 I. 78 12 3.09 I. 74

Several research scientists in post-war Germany The addition of lysine restored the supplementary explored the fortification of bread with milk value of the evaporated milk. products and other protein sources. Bruchner Hutchinson et al. (I 963) studied the protein ( 1949) proposed that up to 40% of the total liquid supplementation of dried bread crumb which used for the preparation of rye bread doughs could provided 85% of the diet and the sole source of be in the form of fresh whey. protein for the rats under test. They report that an Sabiston and Kennedy (I 957) state that white addition of 0.5% L-lysine plus 0.2% L-threonine bread and whole wheat bread containing 0.166% raised the biological quality of British type bread L-lysine (roughly equivalent to the amount of protein into the range which could be ranked as lysine contributed by 6% nonfat dry milk) is "first class". They report that 18% of skim milk roughly equivalent in PER to bread containing powder was necessary to achieve an equivalent 6% nonfat dry milk. The PER of the baked bread, biological value. whether supplemented by nonfat dry milk or its Gates and Kennedy (I 964) compared milk equivalent in lysine, was approximately 20% solids added at 3%, 6% and 12% levels with and lower than the equally supplemented unbaked without 0.25% L-lysine HCI in bread and unbaked ingredients. ingredients. Their results, expressed as adjusted Jahnke and Schuck (I 957) report upon the PERs as determined by the method of Osborne effect of increasing levels of milk solids added et al. (1919) modified by Chapman et al. (1959) with and without additional lysine to (a) bread and are shown in Table I 08 and demonstrate the (b) unbaked ingredients. The results showed a following: (a) an increase in PER with increasing progressive increase in nitrogen efficiency ratio milk solids in both unbaked and baked products; (NER, weight gain in grams per gram nitrogen (b) an average reduction of 21 % in PER when the consumed) with increase in milk solids from 3% ingredients were baked into bread; (c) an overall to 12% in both bread and unbaked ingredients. significant increase in PER with the addition of The addition of 0.25% lysine further increased the 0.25% lysine to milk solids but no significant NER at each level of addition of milk solids. difference between 3%, 6% and 12% milk solids Mauron and Mottu (I 962) compared evapo­ when lysine was present. rated and sweetened condensed milk in iso­ Bains et al. (I 964) compared rat PERs of: nitrogenous quantities in a white flour diet for rats. (A) macaroni made from Indian durum wheat Sweetened condensed milk was found to be semolina, (B) diet (A) fortified with L-lysine HCI, superior to evaporated milk as measured by PER. 0.25 g/100 g protein after processing, (C) enriched HULSE/LAING: PROTEINS OF TRITICALE 147

TABLE 109. Protein content of macaroni, total L-lysine HCI in protein, mean protein intake and PER of plain and enriched macaroni, with and without added lysine HCI, on rat PER (data from Bains et al. 1964).

Protein Total Mean content of L-lysine HCI protein macaroni in protein intake (%) (%) (g) PER

(A) Plain macaroni 12.0 3.16 19.59 1.10 (B) (A)+ L-lysine HCI 0.25 g/100 g 12.0 5.40 26.95 2.11 (C) Enriched macaroni (plus casein and skim milk solids) 17.0 6.80 28.98 2.48 (D) (C) + L-lysine HCI 0.25 g/100 g protein 17.0 9.60 29.08 2.63 macaroni made from durum wheat semolina 90, legume or oilseed protein as a bread protein casein 7, and skim milk solids 3 parts, (D) diet (C) supplement. The recent international interest in fortified with L-lysine HCI, 0.25 g/I 00 g protein, other legumes gives hope that other vegetable after processing. The macaroni was included in proteins might be used to fortify bread in the less 10% protein rat diets fed ad libitum over 28 days. developed countries in the future. The results are given in Table 109. In the developed world soya products pre­ Diet (B) significantly improved the quality of dominate among legumes used as supplements to the protein over diet (A), but fortification with cereal proteins. It is probable that more than 22 milk protein and casein produced a still greater million kilograms find their way into baked improvement. The PER of macaroni, diet (C), products alone. "Soya flours" even in developed was not significantly improved by the addition of countries vary significantly in nature and com­ lysine. position, a fact which causes difficulties in relating It is very well recognized that, from their own data from one source to that from another. resources, developing nations cannot normally The chemical and physical properties of soya seriously consider the supplementation of cereal products vary markedly according to the method foods with milk protein. Nevertheless from time of processing. High fat soya products may vary in to time more affluent nations see fit to present protein content (N x 6.25) from 38 to 46%, low dried milk to those less fortunate. It may be that fat and defatted soya flours from 46 to 53%. in some circumstances the most beneficial use Commercial soybean protein concentrates range of these gifts may be as supplements whereby to in protein from 65% to 70% and soybean isolates enhance the biological value of cereal proteins. from 90 to 95%. The nutritional value also of soya products is influenced by processing conditions. Horan ( 1967) Food Legume and Oilseed Protein compared the protein efficiency (the method is not There is much recorded evidence from ancient stated) of commercial types of defatted soya flours Egypt, Greece and Rome, and from later societies, as illustrated in Table 110. to indicate that bean, pea and other legume flours Data extracted from a variety of sources were added to wheat flour either to extend the including Food and Agriculture Organization available wheat supplies in times of emergency or (I 970b) suggests approximate ranges of biological to provide cheap bread for poor people. McCance values for various kinds of processed soya products and Widdowson (1955) presented a compre­ and these are quoted in Table 111 . hensive historical review of the conflicting opinions Mayr (1948) reported on the addition of 55% concerning the relative nutritional merits of brown protein soya meal to bread in Germany at levels and white breads. They described the supple­ between 2.5 and 5%. At the 2.5% level bread mentation of wheat flour with other cereals and protein content on a dry weight basis increased by legume flours in wartime Europe. I . I% and at the 5% level by 2.2%. Soybean Protein In the past, greater atten- Henry and Kon ( 1949) reported that the com­ tion has been given to soybean than to any other bined effect of 3% skim milk powder and 2.78% 148 MONOGRAPH IDRC-021e

TABLE 110. Effect of heat treatment on protein TABLE 111. PER values of processed soya products quality of commercial defatted soya flours (data from (data from various sources including Food and Horan 1967). Agriculture Organization l970b).

Relative PER Protein (casein= 2.5) Protein Efficiency dispersible (dried skim High fat soya flour 2.0--2.5 Heat treatment index milk=IOO%) Defatted soya flour 2.0--2.5 Soya protein concentrate 2.0--2.5 Negligible heat 90--95 40--50 Soya protein isolate 1.1-2. I Light heat 70--80 50--60 Moderate heat 35-45 75-80 Toasted 8-20 85-90 and 96 at agricultural school K aged I Yz- to 18} years. At school E the flour used was of 87% extraction from imported wheat fortified with 5% soya flour was superior in biological value to 6% of a commercial toasted soya meal. At school K of dry skim milk when included in bread. the flour was of 74% extraction and unfortified. Westerman et al. ( 1954) found no significant Both were supplemented with 2.5 mg riboflavin difference between wheat germ and soya flour and 2.5 g calcium carbonate per kg of flour. The added to enriched wheat flour on the rate of 74% bread contained, on average, per I 00 g growth of rats during the first or second genera­ bread, 8.6 g protein (conversion factor not stated), tion but soya flour added to non-enriched wheat 274 mg lysine and 174 mg methionine. The 81 % flour appeared to promote a faster rate of growth bread contained on average 10.4 g protein, 348 in the second generation than did wheat germ. mg lysine, 186 mg methionine. Other flour products Rats fed enriched flour plus soya flour stored were eaten at both schools, and at school E, the more B vitamins in their livers. traditional white bread and cakes were eaten on Guggenheim and Friedman ( 1960) reported on the Sabbath. bread baked from flours varying in extraction Individual food consumption of the subjects rate from 74 to 95% and fortified with up to 12% was not recorded but flour products provided over soya flour. Lysine (milligrams per gram nitrogen) 40% of the calories in both schools, the experi­ increased with increase in extraction rate and with mental flours providing 32% at school E and 36% percent soya flour added. Threonine increased at school K. There were other differences between and methionine decreased with the percent added the diets at the two schools but at the end of the soya but neither changed appreciably with extrac­ observation period no appreciable differences tion rate. attributable to the bread were found in increase in NPR (Bender and Doell 1957) rose with extrac­ weight and height, change of skin fold thickness, tion rate and with percent ofadded soya. Bornstein general nutritional status and haematological and Lipstein (1962) using the same materials in conditions. feeding trials with chicks, reported a marked Mizrahi et al. (1967) reported on the effect of improvement in growth rate with soya supple­ using isolated soya proteins, produced by an mentation, possibly from the increased dietary isoelectric wash and calcium coagulation, on the protein level and the additional lysine. The growth baking characteristics, acceptance and nutritive rate was more marked than with increasing rate value of bread. They describe how these additions of extraction. The findings of Guggenheim and increased water absorption and 'how loaf volume Friedman and of Bornstein and Lipstein, are in decreased proportionately with the level of protein good agreement. isolate added. The decrease in loaf volume from Later, Guggenheim et al. (1962) reported on mixtures containing less than 6% soya protein feeding trials conducted in Israel over eight was counteracted by the addition of I% of months with adolescent boys and girls consuming lecithin. Protein additions up to 8% of ttfe flour large amounts of bread and doing heavy physical weight did not significantly affect flavour. PER work in addition to their school studies. There of the bread increased with increase in the were 88 boys and girls at agricultural school E percentage of soya protein and was found to be HULSE;LAING: PROTEINS OF TRITICALE 149 m linear relationship with the lysine content of TABLE 112. Effect of additions of pulses with and the bread. without amaranth on rat PER (data from Phansalkar Pomeranz and Finney (1973), United States et al. 1957). Department of Agriculture, have recently been awarded a US Patent No. 3,679,433 covering a PER process for the addition of up to 16% soya flour, or Protein other protein-rich, high-lysine concentrate, with Diet (%) Average S.E. 0.25 to 4.0% glycolipids, to wheat flour to provide (Al Skim milk diet 10.48 2. 57 0.095 acceptable bread containing approximately 70% (B) Wheat 10.58 I. 77 0.082 more protein and three times as much lysine. (C) (B) + bengal gram 11. 11 2. 18· 0.092 The volume of literature covering the supple­ (D) (B) + black gram 10.50 2. 15• 0.100 mentation of wheat and rye with other oilseed and (E) (B) + green gram 10.98 2.22· 0.108 grain legume proteins is sparse compared with (F) (B) +red gram 10.24 2. 19• 0.065 that which treats of soya bean supplements. Some (G) (B) +amaranth 10.94 1.65 0.072 attention has been given to fortification of bread (H) (C) +amaranth 10.94 2.19 8 0.080 with cottonseed flour though a considerably (J) (D) +amaranth 10.32 2. 37• 0.073 (K) (E) +amaranth 10.85 8 greater volume of literature describes the nutri­ 2.23 0.118 (L) (F) +amaranth 10.54 2.35 8 0.080 tional properties of unbaked cereal-cottonseed mixtures. •p < 0.01, all, with the exception of diet (G), Co11onseed Protein When fed to young rats amaranth, significantly different from wheat diet (B). at 10% protein levels, bread containing 10 parts of cottonseed flour per 100 parts of wheat flour produced higher rates of gain per gram of nitrogen gram (chick-pea, Cicer arietinum), black gram consumed than bread without the cottonseed (Phaseolus mungo), green gram (Phaseolus flour (Womack et al. 1954). Womack et al. also radiatus), and red gram ( Cajanus indicus). The compared bread containing skim milk powder and leafy vegetables were agathi (Sesbania grandiflora), bread with skim milk powder and cottonseed amaranth (Amaranthus gangeticus), murungu flour, but found no significant difference in weight (Moringa oleifera) and parpukeerai (Portulaca gain when the two breads were fed isonitro­ olearacea). All materials were locally purchased. genously. However, when fed at the same per­ The diet, about 10% protein (conversion factor centage by weight, the cottonseed flour bread, not stated), consisted of 7% wheat and 3% pulse, having a higher protein content, produced a or 6% wheat plus 3% pulse and 1% amaranth. The significantly higher rate of weight gain. rats were male and female weanling albinos, fed Dalby ( 1969) described how Egyptian Baladi ad libitum and weighed once a week for four weeks. bread was supplemented first with a mixture of Of the leafy vegetables, only amaranth is chick-pea flour and inactive dry yeast and later reported upon. Results are given in Table 112. with two samples of low-gossypol cottonseed There was no significant difference between the flour, the first having been made from a glandless PER, 2.35, of diet (L), (wheat, red gram and cottonseed variety; the second rendered gossypol­ amaranth, protein 10.5%) and the skim milk diet free by solvent extraction. Dalby stated that at (Al. (PER 2.57, protein 10.5%). 7% of the wheat flour, cottonseed flour, contain­ The rat diets contained vitamin mixtures and ing roughly 60% protein, provided a protein since these would not be present in the normal supplementation equivalent to 12% of nonfat milk Indian diet, a trial with bajra (sorghum) plus solids. red gram (pigeon pea) plus amaranth was carried Several other authors have reported upon the out to see if omission of the mixtures would use of various oilseed and legume proteins in affect the PER. The PER was lowered when the bread but most of the published discussion relates salt plus vitamin mixture was not added but the to technological rather than nutritional implica­ difference was not statistically significant. tions of their experiments. It was observed that PER varied according to Other Vegetable Proteins Phansalkar et al. the season in which the experiments were carried ( 1957) added various pulses and leafy vegetables out though no systematic seasonal differences in to wheat in rat diets. The pulses included bengal food intake were observed. 150 MONOGRAPH IDRC-02le

The effect of supplementing a poor Indian diet tain the weight of the subjects. For the first IO based on wheat (PWD) with full fat soya flour, red days on the wheat diet the subjects were in gram dhal (pigeon pea, Cajanus cajan), bengal negative nitrogen balance but for the remainder gram dhal (chick-pea, Cicer arietinum), or skim of the 50-day experimental phase the subjects milk powder, was studied using male albino rats, were in nitrogen equilibrium and displayed good eight rats per group over four weeks, by Daniel health. et al. (1965). The basic PWD consisted of wheat Three blends of hard wheat and bengal gram 78.5 g, red gram dhal 5.0 g, groundnut oil 5.0 g, dahl (Cicer arietinum) obtained in Mysore, India, skim milk powder 0.9 g, common salt 0.3 g, were evalutated in rats by Daniel et al. (1969). The green leafy vegetable (Amaranthus gangeticus) 2.1 blends were (A) 80 wheat, 20 bengal gram (total g, brinjal (egg plant, Solanum melongena) with protein (N x 6.25) 14.3% dry weight basis), (B) potato 8.2 g. This diet provided 11.26% protein 70 wheat, 30 bengal gram (total protein 16. I%), (N determined by micro-Kjeldahl method) and, (C) 40 wheat, 60 bengal gram (total protein 20. 7%). in g/16 g N, 4.5 g lysine and 3.8 g threonine The ratio of wheat to bengal gram protein in the assayed microbiologically. The basic diet (protein blends was (A) 2: I, (B) I: I and (C) I : 3. The 11.26%) was supplemented with bengal gram or lysine, methionine, cystine and threonine contents red gram at a 15% level, soya flour at 5.5% or were assayed microbiologically. The blends were skim milk powder at 9.0% level to provide 2.2% fortified with L-lysine, DL-methionine and DL­ extra protein and corresponding increases in threonine to the levels of these amino acids in lysine and threonine. All these diets were fed with human milk. and without standard mineral and vitamin pre­ The PERs (corrected, Osborne et al. (1919) mixes. taking the PER of skim milk powder as 3.0) were The growth rates of the rats increased signifi­ (A) 2.43, (B) 2.37 and (C) 2.25. The PER of (A) cantly as a result of the supplementation with increased to 2.70 when supplemented with lysine bengal gram or red gram at 15%, or soya flour at and threonine, and to 2. 96 when supplemented with 5%, and they also increased significantly when lysine, threonine and methionine. The PER of vitamins and minerals were added to the basic (B) increased to 2. 72 when fortified with lysine and diet. With both types of supplementation there threonine and to 2.93 with lysine, methionine and was a further significant increase. The PERs of threonine. The PER of (C) increased to 2.66 when the rats fed supplements of bengal gram, red gram fortified with methionine and to 2.80 with and soya to provide 2.2% extra protein were methionine and threonine. The mean weekly gain similar (1.90, 1.92 and 1.93), and when the diets in body weight of the female rats fed blends (A), included vitamins and minerals there was a (B) and (C), with different levels of protein, forti­ significant improvement (to 2.31, 2.41, 2.39) fied with vitamins and minerals were 18.4 g, respectively. There was no significant difference 15.8 g and 17.0 g respectively over a period of 4 between these PERs and those for diets supple­ weeks. Supplementation with lysine and threonine mented with skim milk powder with (2.46) and increased the mean weekly growth rate on blend without (2.46) vitamins and minerals. By the (A) to 23.6 g and on blend (B) to 19.6 g, and criteria employed supplementation of a poor supplementation with methionine and threonine Indian wheat diet with 15% bengal gram (chick­ increased the mean weekly growth rate on blend pea) or red gram (pigeon pea) is roughly equivalent (C) to 20.0 g. to supplementation with 5% soya flour. The results of the PER test and of the growth Bolourchi et al. (1968) examined 12 male test are not strictly comparable but Daniel et al. students aged 19 to 27 years given a diet in which suggest they indicate that a blend of 80 parts of 90 to 95% of the protein came from US commercial wheat to 20 parts of Bengal gram, of about 14.0% enriched wheat flour eaten as bread and rolls protein on a dry basis and fortified with vitamins made without milk. The remainder of the dietary and minerals would meet the protein needs of protein came from fruits and vegetables. The growing children. experimental "core" diet provided 3,320 kcal per Edwards et al. (1971) reviewed earlier work, day, 11.8 g nitrogen, 67.3 protein, 80 g fat and including that of Bolourchi et al. (1968), on the 559 g carbohydrate. All the core diets were nutritive value of wheat for adult men. They also supplemented with protein-free calories to main- investigated the nutritional value of wheat flour HULSE/LAING: PROTEINS OF TRITICALE 151

protem m 12 male students, aged 23 to 30 years growth response to be proportional to the lysine during a 74-day period. The commercial white added from the dry yeast and dry milk supple­ flour was fed in the form of bread made without ments. either milk or any other source of animal protein. McCollum (1945) reported on the nutritional The wheat diet contained 46 g protein, of which benefit of adding dried brewers' yeast and indi­ 35 g was provided by the bread and 11 g by such cated that palatability was not seriously impaired. other plant foods as potatoes, turnips, green Nevertheless, both he and Schwarz et al. (1942) vegetables and fruit. In other diets 20% of the reported a significant decrease in bread quality, wheat nitrogen provided was replaced, isoni­ assessed technologically, as the amount of added trogenously, by pinto beans, rice, or peanut butter. dry yeast was increased. The daily calories supplied were: wheat diet Sure (1948) attributed a superior weight gain in 3,036; wheat plus pinto bean 3,006; wheat plus rats fed wheat flour enriched with dried food yeast rice 3,043; wheat plus peanut butter 3,043. to the lysine added by the yeast. Nitrogen balance, blood urea nitrogen, urinary Seeley et al. (1950) found that when non-viable urea, plasma lipid concentration, and essential dried yeast was added to bread, at a concentration and non-essential amino acids were determined at not exceeding 3%, rats fed on the resulting bread the beginning and end of four 15-day intervals showed an increase in daily weight gain over following the wheat diet regimen. controls. Dried yeast was found to be, nutri­ Nitrogen balance was maintained over the trial tionally, a better supplement than nonfat dry milk period and Edwards et al. conclude that a diet but the maximum increase in weight gain occurred providing 46 g protein per day of which 76% in bread containing both dried yeast and nonfat of the nitrogen was supplied by wheat and the milk solids. remainder by potatoes, other vegetables and fruit Lang (1950) discussed the fortification of bread is adequate for nitrogen maintenance in adult with lysine and torula yeast and emphasized that man. Replacement of 20% of the nitrogen with a greater improvement in biological value was pinto beans, rice or peanut butter did not signifi­ obtained than by using whole wheat flour. cantly improve the utilization of the wheat proteins. Cremer et al. (1953) discussed various means of supplementing bread with protein, including milk Protein from Microorganisms powder and dried yeast, and stated that the addi­ Yeast acts in bread doughs as a leavening agent tion of 3% protein significantly raised the bio­ and, until recently, yeast has been studied nutri­ logical value of the bread. tionally more as a source of B-complex vitamins Hundley et al. (l 956) described the fortification than of protein. The natural yeast cell contains of bread with two types of algae: (i) Scenedesmus about 30% dry matter of which more than 50% obliquus (ii) Ch/ore/la pyrenoidasa. The first con­ is crude protein (N x 6.25). Its chemical composi­ tributed significant amounts of both lysine and tion is comparatively well known (Von Loesecke threonine; the second, though an adequate source 1946, Inskeep et al. 1951) but some uncertainty is of threonine, was comparatively deficient in lysine. apparent concerning the long term effects of Pomeranz (1962) found that the addition of feeding large amounts of yeast to human beings. 3%, 5% and I 0% of dry yeast to bread raised the Those yeasts which over thousands of years have initial protein content from 13.8% to 14.4%, been consumed in bread, beer and wine have been 14.7% and 15.9% respectively. By microbiological ingested in relatively small daily quantities. assay he determined that the control lysine content During World War II workers, mostly in of0.46% was raised respectively to 0.49, 0.51 and Germany, examined the influence of dried yeast 0.59%. The protein (N x 6.25) and lysine content in bread. Schwarz et al. (l 942) found that addi­ of the active dried yeast were 42.4% and 2.97% tions of 2.5% and 5% of dried brewers' yeast respectively. significantly improved the vitamin B-complex and Spicer (1970, personal communication) under­ protein content of white bread. took the screening of a large number of micro­ Light and Frey (1943) reported that 5% dry organisms found in carbohydrate sources and yeast containing approximately 50% protein com­ describes the discovery of a microfungus with a pared favourably with bread containing 6% protein content of 45% and NPU in the region of nonfat milk solids. These authors found the 70, which grows well on a variety of hydrolyzed 152 MONOGRAPH IDRC-021e

TABLE 113. Lysine, threonine, methionine and cystine TABLE 114. Effect of addition to poor wheat diets of content with and without petroleum yeast (in grams/ 6% yeast on protein content of diet and rat PER (data 16 g N), and protein score of rat diets (data from from Narayanaswamy et al. 1972). Narayanaswamy et al. 1972). Protein content Poor Diet (dry basis,%) PER wheat Petroleum diet PWD plus (A) Poor wheat diet 12.46 1. 50 yeast (PWD) 6% yeast (B) (A)+ mineral and vitamin premix 12.38 2.02 Lysine 7.4 3.2 3.8 (C) (A)+ 6% yeast 14.48 1. 86 Threonine 5.3 3 .1 3.5 Methionine 1.4 I. 9 1 .8 (D) (C) +mineral and Cystine 0.5 I. 2 I. 1 vitamin premix 14.36 2.34 Protein Score 34 50 52 (assuming hen's egg as 100) in PER. The differences in PERs of diets (A) 1.50, (B) 2.02, (C) 1.86 and (D) 2.34, were all significant. Yanez et al. ( 1973) found that the substitution starches and the sucrose present in cane juice and of Chilean flour (70% extraction rate, Florana molasses. The organism is being studied as a wheat) by Candida utilis, cultivated on sugar beet protein supplement in bread and other cereal molasses, at levels of 0, 3, 6 and I 0'%,, improved foods. Spicer does not indicate the level of the crude protein (N x 6.25) content of the digestibility nor the content of nucleic acids resultant bread from 14.4% at the zero level to present. 21.3% at the 10% level. The rat PER (Chapman Narayanaswamy et al. ( 1972) compared the et al. 1959) increased significantly from 0.84 at PER for young female rats, eight per group, fed zero level to 1.14 (3/~ level) to 1.45 (6% level) to ad libitum over 28 days, of a poor wheat diet 1.74 (I0'/0 level) but did not attain the PER of supplemented by yeast grown on hydrocarbons. 2.71 of the casein diet. Loaf volume decreased The yeast was composed of, in grams per 100 g, noticeably and colour darkened as fortification moisture 9.8, protein (N x 6.25) 45, fat I. I. levels rose above I%. Yanez et al. point out that mineral matter I 0.3, calcium 0.16, phosphorous when the nucleic acids of yeast are metabolized by 2.10 (Association of Official Agricultural Chemists human beings the final product is uric acid, which 1955). in excess may lead in susceptible subjects to gout The experimental diets consisted of a basic or the formation of stones in the urinary tract. mixture of vegetables, peanut oil, skim milk Food and Agriculture Organization ( 1971) have powder and common salt totalling 16.59 to set the maximum tolerable intake of nucleic acids which were added: Diet (A) 78.5 g wheat, 5.0 g at 2 gperday, equivalent to I 0 to 30 g of unicellular bengal gram; diet (B) 75.5 g wheat, 5.0 g bengal proteins. In Chile bread enriched with Candida gram, 2.0 mineral mix, 1.0 vitamin premix; diet utilis at the 3, 6 and I 0% levels would provide, (C) 72.5 g wheat, 5.0 g bengal gram, 6.0 g petro­ respectively 400, 800 and 1300 mg nucleic acids leum yeast; diet (D) 69.5 g wheat, 5.0 g bengal daily, assuming an average consumption of 250 g gram, 6.0 g petroleum yeast, 2.0 g mineral mix, bread daily per child. This level is much lower 1.08 vitamin premix. than the recommended maximum. The lysine, threonine, methionine and cystine contents (assayed microbiologically) of the diets is Fish Proteins shown in Table 113. The protein content (micro­ Before presenting their review of supplementa­ Kjeldahl N, conversion factor unstated) and the tion with fish proteins the authors would offer a PERs are given in Table 114. word of caution to the reader. The terms "fish

The addition of 6% yeast, providing about 2/0 flour", "fish protein concentrate", "marine pro­ extra protein in the diets, plus vitamins and tein concentrate" appear in the literature some­ minerals, produced highly significant increases in times to designate closely similar materials, some­ the rate of rat growth, and significant improvement times to describe widely different products. In HULSE/LAING: PROTEINS OF TRITICALE 153

TABLE 115. Effect of supplementing wheat flour diet with fish flour on rat PER (data from Metta 1960).

Total Percent Dietary food Weight of protein intake gain Diet diet (%) (g) (g) PER

(A) Whole wheat flour 66 12.9 280 71±1.5 1.980±0.038 (B) Whole wheat flour 65 13.6 280 81±I.3' 2.127±0.035 3 fish flour (C) Whole wheat flour 63 14.8 280 102± 1.1' 2.470±0.028' fiish flour 3 (D) Enriched wheat flour 63 9.7 279 32±3.1 1.183±0.096 cellulose 3 (E) Enriched wheat flour 62 fish flour 10.6 279 56±4. Jb l .882±0.134b cellulose 3 (F) Enriched wheat flour 60 fish flour 3 12. I 279 79±2.8' 2. 364 ±0 .077b cellulose 3

•0.01

(B), diet (A), plus fish flour replacing 1/ 0 of the plus fish flour, 2 lb/I 00 lb wheat flour; (C), diet wheat flour; (C), diet (A), plus fish flour replacing (A) plus fish flour, 4 lb/l 00 lb wheat flour; (0), 3% of the wheat flour; (0), basal diet, plus en­ diet (A) plus fish flour, 6 lb/100 lb wheat flour. riched wheat flour providing 63% of the diet with The results are presented in Table 116. PER in­ 3% cellulose; (E), basal diet, plus enriched wheat creased with increasing addition of fish flour. flour providing 62% of the diet, fish flour 1% and In a second trial the diets, also fed at the 10% cellulose 3%; (F), basal diet, plus enriched wheat protein level, included: (E) casein; (F) enriched flour providing 60% of the diet, fish flour 3% white bread made with water; (G), as (F) with 10% and cellulose 3%. The fish flour contained 83.1 % fish flour replacing an equivalent amount of white protein (N x 6.25). flour; (H) enriched white bread including 4.2% 154 MONOGRAPH IDRC-021e

TABLE 116. Effect of supplementation of whole wheat TABLE 118. Effect of defatted fish flour on lysine flour with defatted fish flour on rat PER (data from contentofbreadand flour proteins (data from Morrison Morrison and Campbell 1960). and Campbell 1960).

Diets Lysine content (g/16 g N) (A) (B) (C) (D) Whole wheat flour 3.02 Fish flour added + 2 parts fish flour 3.63 lb/ I 00 lb wheat flour 0 2 4 6 + 4 parts fish flour 4.37 Weight gain, g 37 67 84 102 + 6 parts fish flour 4.68 Food consumption, g 249 314 337 365 PER 1.49 2.12 2.48 2.80 White bread 2.43 + 10% fish flour 6.32 Milk bread (4.2% milk solids) 3.16 milk solids; (J), as (H) with I 0% fish flour replac­ + 10% fish flour 5.86 ing an equivalent amount of white flour. The results on rat PER are shown in Table 117. The lysine contents of the flour and bread diets, lb wheat flour, as in the first trial, increased the determined microbiologically, are shown in Table protein content of the mixture by 10% but in­ 118. creased the "complete" protein content by 42%. The bread used in diet (G) had a protein content Fish flour contains approximately I 0.9 g lysine per of 16.5% and that used in diet (J) a protein content 16.0 g N (Rogers et al. 1959) so it also contributes of 18%, air-dry basis. These "fish-flour" breads "extra" lysine. showed reduced loaf volume, were darker in The protein rating (Campbell and Chapman colour, and somewhat "rubbery" in texture 1959) of the white bread (diet F) was 11.1 and of but the smell and taste were not noticeably affected. the bread containing fish flour (diet G) was 51.5. A highly significant correlation of 0.993 was Jansen et al. (1966) studied the effect ofalcohol­ found when the lysine content was plotted against extracted fish flour at 3% and 6% in combination the PER, indicating that the PER values in these with L-lysine and L-lysine plus DL-threonine, the diets are a direct function of the percentage additions being made to standard white bread lysine in the protein. Morrison and Campbell after drying. From rat feeding studies, they con­ cite Rogers et al. ( 1959) and Flodin ( 1959) as also cluded that the addition of lysine and threonine observing a significant relationship between lysine doubled the utilizable bread protein, with lysine concentration and PER in foods low in lysine. alone being responsible for approximately 75% of Howard et al. ( 1958) has suggested that an the increase. Bread supplemented with 6% of ideally balanced "complete" protein should con­ fish flour displayed a protein retention efficiency tain 5.3 g lysine per 16.0 g N. The supplemental of 58; the addition of 0.2% of lysine raised this value of fish flour, based on its content of "com­ value to 72; 0.4% lysine plus 0.15% threonine with plete" protein, was calculated according to the no fish flour gave a value of 78 compared to 88 formula given by Howard et al. (1958) and found for the reference sample of casein. Since these to be 185. The addition of2 lb offish flour per 100 additions were made to dry bread after baking,

TABLE 117. Effect of supplementation of white bread with defatted fish flour on rat PER (data from Morrison and Campbell 1960).

(J) (G) (H) Milk bread+ (E) (F) Bread+ 10% Milk 10% fish Casein Bread fish flour bread flour

Weight gain, g 109 23 145 43 130 Food consumption, g 311 177 376 216 363 PER 3.49 1.30 3.87 1.97 3.59 HULSE/LAING: PROTEINS OF TRITICALE 155

TABLE 119. Biological quality of bread and of bread with 6% fish flour, 12% skim milk powder and 6% fish flour plus 0.5% L-lysine (data from Yanez et al. 1967).

Protein calories Net protein (%) concentration NPU,. Score

Bread 35.4 10.4 3.7 35.4 45 + 6% fish flour 42. 8 14.3 6.1 48.9 72 + 12% skim milk powder 44. 5 13.5 6.0 50.4 71 + 6% fish flour + 0.5% L-lysine 58. 8 14.6 8.6 71.4 70

they do not reflect any degree of loss which might FPC. Sidwell states that as the FPC increased the occur during baking. degree of sweetness decreased and the colour Sidwell (1967) discussed the nutritional ad­ changed from bright yellow to dull grey yellow. vantages of supplementing wheat flour with fish Yanez et al. (196 7) examined samples of fish protein concentrate as developed and produced flour produced in a pilot plant at Quintero, Chile by the Bureau of Commercial Fisheries in the from Merluccius gayi. In the products examined, United States. He stated that a mixture of 10% available lysine (Carpenter 1960) ranged from FPC and 90% wheat flour in cookies showed a 8.3 to 9.5 g per 100 g of crude protein (Viobin PER of 3.0 compared to 3.4 for casein. The FPC Corp. product, for comparison, 8. 7), and NPU was used as an ingredient of bread and cookies. (Miller and Bender 1955) from 63.5 to 70.7 FPC was added to the bread formula at levels of (Viobin 69.2). Fish flour was added to 5, 10, 15, 20 and 25%. At all levels of addition "marraquata" bread at the 10% level and com­ there was a noticeable effect on crumb colour and pared with control bread, and bread with 12% loaf volume, the former becoming progressively skim milk powder. The protein content in grams darker and the latter smaller with progressive per 100 g dry matter was: control bread 10.2 increases in FPC. The crude protein content of the (N x 5. 7), bread with 6% fish flour 13.9 (N x 5.9), bread (N x 6.25 moisture free basis) rose from bread with 12% skim milk powder 13.2 (N x 5.9). 15.0% in the control to 33.9% at a 25% level of The biological quality of the breads is shown in addition. The author's estimates of protein and Table 119. It is not clear from the text exactly how lysine in a 40 g slice of bread were: these results were obtained. % FPC 0 5 10 15 20 25 'Tallanines", a pasta product, was made locally, Protein, g 3. 8 4. 6 5. 7 6. 4 7 .1 7 .8 under normal manufacturing conditions, con­ Lysine, mg 70 96 136 154 214 259 taining 10% of fish flour. The chemical composi­ The crude protein content of cookies increased tion and the biological value are shown in Table from 5.4% at zero addition to 9.8% at 15% of 120.

TABLE 120. Moisture, protein content and biological quality of commercial pasta, pasta enriched with 10% fish flour, and the unprocessed ingredients of the enriched pasta (data from Yanez et al. 1967).

Net Protein Protein protein Moisture (N x 6.25) calories concen-

(%) (%) NPU0 • (%) tration NPU,, Score

Commercial pasta 11. 5 10.2 38.4 11.9 4.6 40.2 47 Pasta enriched with 10% fish flour 10.6 16. I 47.0 19. I 9.0 64.4 73 Ingredients of enriched pasta 14.1 15.8 51.6 19.2 9.9 71. 8 73 156 MONOGRAPH IDRC-02le

TABLE 121. Biological quality of roasted whole wheat meal without and with 10% fish flour (data from Yanez et al. 1967).

Protein calories Net protein NPUOP (%) concentration NPU,. Score

Roasted whole wheat meal 41. 2 8.2 3.6 41.2 47 Roasted whole wheat meal with 10% fish flour 70.2 11.6 8.2 81.6 72

The biological quality of roasted whole wheat pared from red hake ( Urophycis chuss) by extrac­ meal, enriched with I 0% fish flour is given in tion with isopropyl alcohol. Table 121. In summary, when unprocessed wheat flour "Ulpo", a preparation of roasted whole wheat mixtures were fed at I 0% protein level, maximum meal with water or milk, is widely eaten by young weight gain, PER, NPU and total protein and fat children, especially in rural areas of Chile. A in the carcasses were produced by 15% FPC and ration of "ulpo" consisting of a mixture of roasted 0.2 to 0.4% lysine. Maximum responses from FPC whole wheat meal 70 g, fish flour I 0 g and sugar were greater than those from lysine. When bread 20 g, mixed with 200 ml of cold water was pre­ baked from the wheat flour mixtures was fed at sented to each of 300 children aged 9 to 14 years the same protein level, 25% FPC and 0.4% lysine and found completely acceptable. produced maximum weight gain and PER. Protein When fish protein concentrate was used as a ratings for bread supplemented with 10 to 25% supplement to wheat in the diet of Peruvian FPC were 35 to 150% higher than those for bread children, acceptance and consumption were excel­ supplemented with lysine. When bread was in­ lent during the six years of the project (Baertl cluded in diets on the basis of 80% by weight, I 0% 1971 ). The study was carried out in four villages FPC or 0.4% lysine produced the highest weight each of about 1000 inhabitants where evidence of gains, the weight gain being 36% higher with FPC poor growth, a high incidence of malnutrition than with lysine. and mortality existed among pre-school children. A variety of authors including Donoso et al. Villages I and 2 served as controls; in village 3 (1963), Santa Maria (1969), Bacigalupo (1969) each family received enough ordinary wheat describe the addition of dehydrated fish flour noodles to provide a daily supplement of 250 kcal (harina de pescado) at various levels up to 12% and 7 g of wheat protein per person. In village 4, of the wheat flour but their publications are con­ fish protein concentrate replaced I 0% of the noodle cerned more with acceptability than with improve­ flour. There were only minor effects on growth but ment in nutritional value. the mortality of infants and pre-school children Sen et al. (1969) describe several alternative in villages 3 and 4 dropped in comparison with methods of producing fish protein concentrate the preceding 12-year pattern and was not from Bombay duck (Harpodonn nehereus) and the matched in villages I and 2. It is worthy of note influence of processing conditions upon the that during the experimental period, improve­ nutritional and organoleptic properties. The ments in housing and sanitation took place in samples were incorporated into bread, chapatis, villages I, 2 and 3 but not in village 4. puris and other cereal foods. Stillings et al. (1971) compared the PER and NPU of wheat flour, and of bread made from it, Conclusion unsupplemented and supplemented with either Though the most widespread benefit to man­ fish protein concentrate (FPC) or lysine. The kind would probably result from the development samples were commercial (United States) baker's of nutritionally superior varieties of triticale, the grade wheat flour of 86.9% dry matter, protein biological value of products derived from triticale, 14% (N x 6.25), to which was added L-lysine HCI wheat and rye can be improved or impaired by or FPC of 94.9% dry matter, protein 85%, pre- how they are processed. Among malnourished HULSE/LAING: PROTEINS OF TRITICALE 157 peoples it is important that as much as possible of oilseeds grown in the triticale producing areas. the protein-rich aleurone layer be retained in the Milling devices which can process both cereal milled flour. More efficient milling techniques grains and legumes within the same facility will, applicable in the developing countries whereby it is believed, make possible the production and to retain the aleurone layer in the flour should use of nutritious cereal-legume mixtures. The therefore be sought. In addition research is needed International Development Research Centre is wherewith to produce inexpensive acceptable encouraging a series of integrated projects with protein supplements derived from legumes and this end in view. Epilogue

Triticale is well along the way to becoming the first commercially viable cereal grain to result from a scientifically controlled genetic wide-cross. Several triticales produce significantly higher yields of grain than the most productive known bread wheats grown under comparable conditions. Lysine contents equivalent to high lysine maize, and protein contents in the same order as bread wheats are also to be found among advanced triticale lines. More research is required to determine the overall biological value of advanced triticales in animal feeds, much of the earlier reported work being inconclusive. Greater attention should also be given to exploring fully the potential of the rye parent in the synthesis of triticales of higher biological value. Much progress is apparent in the improvement of inherent fertility and in kernel characteristics. Grain and endosperm shrivelling, though not yet totally eliminated, is currently of a significantly lower incidence than in the past. Further improvements in genetic, agronomic, technological and biological properties may be confidently expected. These are the tangible and predictable products of the Triticale Project. The Triticale Project provides more than these tangible benefits. The Project clearly demonstrates the feasibility of producing viable plants from wide genetic crosses. The Project also serves as a model of what can be achieved by a scientifically integrated approach to plant breeding in which equal concern is afforded to the plant's agronomic properties, its biological quality and its utility. The Triticale Project demonstrates that, in the words of Radhakrishnan, "We can take a hand in shaping the future of things."

158 References

ABROL, Y. P., D. c. UPRETY, V. P. AHUJA, AND M. S. ANDERSON, R. A., A. C. STRINGFELLOW, -AND E. L. NAIK. 1971. Soil fertilizer levels and protein GRIFFIN, JR. 1972. Preliminary processing studies quality of wheat grains. Aust. J. Agric. Res. 22(2): reveal triticale properties. Northwest. Miller. 279 195-200. (2): 10--13. ADOLPH, W. H., AND Y-F. TSUI. 1935. Effect of ASSOCIATION OF FOOD SCIENTISTS AND TECHNOLOGISTS steaming compared to baking on the nutritive value (INDIA). 1969. Protein fortification of foods. Pro­ of wheat bread. Chin. J. Physiol. 9: 275-284. ceedings of a seminar at Jadavpur University, ADRIAN, J., AND R. FRANGNE. 1969. Nutritional Calcutta, India, 15-16 February, 1969. Association value of biscuits supplemented with pure lysine or of Food Scientists and Technologists (India), 92/1, with various protein sources: groundnut, fish or Alipore Road, Calcutta-27. 126 p. milk (in French). Ind. Aliment. Agric. 86: 801-806. ASSOCIATION OF OFFICIAL AG RI CULTURAL CHEMISTS. AKESON, W. R., AND M. A. STAHMANN. 1964. A 1950. Official methods of analysis. 7th ed. Associa­ pepsin pancreatin digest index of protein quality tion of Official Agricultural Chemists, Washington, evaluation. J. Nutr. 83: 257-261. D.C. ALLEE, G. L., AND R.H. HINES. 1972a. Nutritional 1955. Official methods of analysis. 8th ed. adequacy of triticale for finishing swine. J. Anim. Association of Official Agricultural Chemists, Washington, D.C. Sci. 35(5): 1101. I 972b. Nutritional value of triticale for growing 1960. Official methods of analysis. 9th ed. swine. J. Anim. Sci. 35(5): 1101. Association of Official Agricultural Chemists, Washington, D.C. ALLISON, J.B. 1959. The efficiency of utilization of 1965. Official methods of analysis. 10th ed. dietary proteins, pp. 97-116. In A. A. Albanese [ed.] Protein and Amino Acid Nutrition. Academic Press, Association of Official Agricultural Chemists, Washington, D.C. New York. ASSOCIATION OF OFFICIAL ANALYTICAL CHEMISTS. 1964. The nutritive value of dietary proteins, 1970. Official methods of analysis, 11th ed. pp. 48-86. In H. N. Munro and J. B. Allison [ed.] Association of Official Analytical Chemists, Mammalian Protein Metabolism. Vol. 2. Academic Washington, D.C. Press, New York. ATTIA, F., AND R. D. CREEK. 1965. Studies on raw ALTSCHUL, A. M. 1965. Proteins-their chemistry and heated wheat germ for young chicks. Cereal and politics. Basic Books, New York. Chem. 42(5): 494-497. ALTSCHUL, A. M., AND D. ROSENFIELD. 1970. Pro­ AUSTIN, A., V. K. HANSLAS, H. D. SINGH, AND P. tein Supplementation: satisfying man's food needs. RAGHA VIAH. 1970. Protein survey of improved Progress 54: 76--84. Indian wheat (Triticum aestivum L.) varieties. AMERICAN ASSOCIATION OF CEREAL CHEMISTS. 1957. Indian J. Agric. Sci. 40(4): 302-308. Cereal Laboratory Methods. Minneapolis, Min­ AUSTIN, A., AND R. K. M1R1. 1961. Effect of nitro­ nesota. gen and irrigation on the protein and gluten content 1962. AACC Approved methods (Cereal of some New Pusa wheats. Indian J. Plant Physiol. Laboratory Methods). 7th ed. American Association 4:150--155. of Cereal Chemists, St. Paul, Minnesota, 55104. AXFORD, D. W. E., N. CHAMBERLAIN, T. H. COLLINS, 1968. Approved methods. Crude protein-Udy AND G. A. H. ELTON. 1963. The Chorleywood dye method AACC method 46--14. (Issued May Process. Cereal Sci. To-day 8: 265. 1968). 3340 Pilot Knob Road, St. Paul, Minnesota AYKROYD, W. R., AND J. DOUGHTY. 1970. Wheat 55121. in human nutrition. FAO Nutr. Studies No. 23. ANDERSON, G. L. 1963. Effect of iron/phosphorus Food and Agriculture Organization of the United ratio and acid concentration on the precipitation of Nations, Rome. ferric inositol hexaphosphate. J. Sci. Food Agric. BACIGALUPO, A. 1969. Protein-rich cereal foods in 14: 352-359. Peru. pp. 288-304. In Max Milner [ed.] Protein­ ANDERSON, R. A. 1963. Wet milling properties of enriched Foods for World Needs. American Associa­ grain: bench-scale study. Cereal Sci. Today 8(6): tion of Cereal Chemists, St. Paul, Minnesota 190--192, 195-221. 55104.

159 160 MONOGRAPH IDRC-02le

BAER TL, J. M. 1971. Study of the effect of dietary BEECH, D. F., AND M. J. T. NORMAN. 1966. The supplementation on Peruvian children, pp. 294-297. effect of time of planting on yield attributes of wheat In N. S. Scrimshaw and A. M. Altschul [ed.] Amino varieties in the Ord River Valley. Aust. J. Exp. acid fortification of protein foods. Rept. of an Int. Agric. Anim. Hush. 6: 183-192. Conf., Massachusetts Institute of Technology, BENDER, A. E. 1957. Nutritive value of bread Cambridge, Mass. 1969. ISBN 0 262 19091. 5. protein fortified with amino acids. Proc. Nutr. Soc. BAINS, G. S. 1953. Effect of commercial fertilizers 16(2): xviii-xix. and green manure on yield and nutritive value of 1965. The balancing of amino acid mixtures and wheat. 11. Nutritive value with respect to general proteins. Proc. Nutr. Soc. 24(2): 190-196. composition, thiamine, nicotinic acid and the bio­ 1966. Nutritional effects of food processing. logical value of the protein of the grain. Cereal J. Food Technol. 1(4): 261-289. Chem. 30(2): 139-145. 1969. Newer methods of assessing protein BAINS, G. S., S. VENKATRAO, D. S. BHATIA, AND V. quality. Chem. Ind. July 5, 904-909. SUBRAHMANYAN. 1964. Effect of lysine and milk­ 1973. Chemical scores and availability of amino protein supplements on the protein efficiency ratio acids, pp. 167-177. In J. W. G. Porter and B. A. of wheat macaroni. Br. J. Nutr. 18: 241-244. Rolls [ed.] Protein in human nutrition, Academic BAKER, R. J., K. H. TIPPLES, AND A. B. CAMPBELL. Press, London. 1971. Heritabilities of and correlations among BENDER, A. E., AND B. H. DOELL. 1957. Biological quality traits in wheat. Can. J. Plant Sci. 51(6): evaluation of proteins: a new aspect. Br. J. Nutr. 441-448. 11(2): 140-148. BALLESTER, D., M.A. TAGLE, AND G. DONOSO. 1962. BENDER, A. E., AND D. S. MILLER. 1953. A new Utilization of the protein of wheat, maize and some brief method of estimating net protein value. derivatives in popular consumption (in Spanish). Biochem. J. 53: vii. Nutr. Bromatol. Toxicol. 4: 235-243. BENDICENTI, A., M. BOGILIOLO, P. MONTENERO, AND BANNERJEE, R. M., AND N. B. DAS. 1964. Nutri­ M. A. SPADONI. 1957. Amino acid content of tive value of some pure strains of Indian wheat. some varieties of Italian wheat of varying protein Indian J. Exp. Biol. 2: 114-116. contents (in Italian). Quad. Nutr. 17: 149-158. BARAKAT, M.A., S. I. FAKHRY, AND M.A. M. KHALIL. BENNION, E. B., AND G. S. T. BAMFORD. 1973. 1970. Interaction effects of salinity of the soil and The technology of cake making. 5th ed. Leonard nitrogen fertilizer on wheat yield of grain and Hill, London. 432 p. protein. U.A.R.J. Soil Sci. 10(2): 169-186. BERNHART, F. W., AND R. M. TOMARELLI. 1966. A BARNESS, L.A., R. KAYE, AND A. VALYASEVI. 1961. salt mixture supplying the National Research Lysine and potassium supplementation of wheat Council estimates of the numerical requirements of protein. Am. J. Clin. Nutr. 9: 331-344. the rat. J. Nutr. 89: 495-500. BARTNIK, J. 1960. Effect of the extraction rate on the BERRY, c. P., B. L. D'APPOLONIA, AND K. A. GILLES. digestibility of milling products of rye (in German). 1971. The characterization of triticale starch and its Nahrung. 4(1): 42-51. comparison with starches of rye, durum, and HRS 1964. Investigations into the nutritive value of wheat. Cereal Chem. 48(4): 415-427. the proteins of rye flour of different extraction rates BIXLER, E., P. J. SCHAIBLE, ANDS. BANDEMER. 1968. (in German). Nahrung. 8(6): 511-518. Preliminary studies on the nutritive value of triti­ BARTON-WRIGHT, E. c. 1952. The microbiological cale as chicken feed. Quart. Bull. Michigan Agric. assay of the vitamin B-complex and amino acids. Exp. Stn. 50(3): 276--280. Sir Isaac Pitman, London. 189 p. BLACKBURN, S. 1968. Amino Acid Determination BAYZER, H., AND H. H. MAYR. 1967. Influencing -Methods and Techniques. Marcel Dekker, New the amino acid composition of rye proteins by York. nitrogen fertilization and chlorcholine chloride (in BLOCK, R. J., P. R. CANNON, R. W. WISSLER, C. H. German). Z. Lebensm. Untersuch.-Forsch. 133(3): STEFFEE, JR., R. L. STRAUBE, L. E. FRAZIER, AND 215-217. R. L. WOOLRIDGE. 1946. The effects of baking and BEARE-ROGERS, J. L., AND E. A. NERA. 1972. toasting on the nutritional value of proteins. Arch. Cardiac lipids in rats and gerbils fed oils containing Biochem. 10: 295-301.

C22 fatty acids. Lipids. 7: 548-552. BLOCK, R. J., AND H. H. MITCHELL. 1946 The BEAUDOIN, R., J. MAYER, AND F. J. STAVE. 1951. correlation of the amino acid composition of proteins Improvement of protein quality of whole wheat. with their nutritive value. Nutr. Abstr. Re~. 16(2): Proc. Soc. Exp. Biol. Med. 78: 450--451. 249-278. BECCARI, G. 1745. De Bononieuse scientiarum et BLOCK, R. J., AND K. w. WEISS. 1956. The amino Artium lnstituto Atque Academia. 2(1): 122-127. acid handbook. C. C. Thomas. Springfield, Illinois. REFERENCES 161

BoAs FIXEN, M.A., AND H. M. JACKSON. 1932. The BRESSANI, R., F. VITERI, AND L. G. ELIAS. 1973. biological values of proteins. IV. The biological The usefulness of results with human beings in values of the proteins of wheat, maize and milk. standardizing rat assays, pp. 293-316. In 1. W ..G. Biochem. J. 26: 1923-1933. Porter and B. A. Rolls [ed.] Proteins in human BOLOURCHI, S., C. M. FRIEDEMANN, AND 0. MICKELSEN. nutrition. Academic Press, London. 1968. Wheat flour as a source of protein for adult BRESSANI, R., D. WILSON, M. BEHAR, M. CHUNG, human subjects. Am. 1. Clin. Nutr. 21(8): 827-835. AND N. s. SCRIMSHAW. 1963. Supplementation Boorn, R. G., AND T. MORAN. 1946. Digestibility of cereal proteins with amino acids. IV. Lysine of high-extraction wheat in flours. Lancet. 251 : supplementation of wheat flour fed to young 119. children at different levels of protein intake in the BORLAUG, N. E. 1973. Civilization's Future. A call presence and absence of other amino acids. J. Nutr. for international granaries. Sci. Public Affairs. 79(3): 333-339. 29(8): 7-15. BRESSANI, R., D. L. WILSON, M. BEHAR AND N. S. BORNSTEIN, S., AND B. LIPSTEIN. 1962. Evaluation SCRIMSHAW. 1960. Supplementation of cereal of the nutritive quality of different types of bread proteins with amino acids. III. Effect of amino acid by feeding trials with chicks. Israel J. Agric. Res. supplementation of wheat flour as measured by 12(2): 63-74. nitrogen retention in children. J. Nutr. 70: 176--186. BORONOEVA, G. S. AND E. D. KAZAKOV. 1969. BRIGGLE, L. W. 1969. Triticale-a review. Crop Amino acid composition of spring rye grain (in Sci. 9(2): 197-202. Russian and English). Prikl. Biochim. Mikrobiol. BROWN, J. W., N. W. FLODIN, E. H. GRAY, AND 0. E. 5(3): 314-317. PAYNTER. 1959. The effect of lysine supplementa­ BOWLAND, J.P. 1968. Triticale as a grain for market tion on the protein efficiency of high protein breads. pigs. 47th Annual Feeders' Day Rep. Dept. of Cereal Chem. 36(6): 545-553. Animal Science, University of Alberta, Edmonton. BRUCHNER,-. 1949. The protein problem in nutrition BOYNE, A. W., S. A. PRICE, G. D. ROSEN, AND J. A. and the food industry (in German). Getreide STOTT. 1967. Protein quality of feeding stuffs: 4. Mehl Brot. 2: 2-5. Progress report on collaborative studies on the BRUGGEMANN, J., AND H. ERBERSDOBLER. 1967. The microbiological assay of available amino acids. Br. amino acid content of wheat by-products and wheat 1. Nutr. 21(1): 181-206. germ (in German). z. Tierphysiol. Tierernaehr. BRADLEY, W. B. 1965. Bread as a Food. Northwest Futtermittelkd. 22: 173-181. Miller, 272(4): 17-19. BRUNE, H., AND E. THIER. I 968a. Variability of BRAGG, D. B. 1969. Poultry Sci. 48: 2135-2137. biological value of protein of different types of BRAGG, D. B., D. A. GUY, D. E. GREENE, P. w. cereal (variants with manuring and site). I. Method WALDROUP, AND E. L. STEFANSON. 1966. Com­ of estimating differences in quality with pigs (in parison of amino acids in milo and corn. Poultry German). Z. Tierphysiol. Tierernaehr. Futtermit­ Sci. 45: I 072. telkd. 24: 40-49. BRAGG, D. B., C. A. IVY, AND E. L. STEPHENSON. 1969. I 968b. 2. Description of samples and results Methods for determining amino acid availability of ofanalysis (in German). z. Tierphysiol. Tiererenaehr. feed protein. Poult. Sci. 48: 2135-2137. Futtermittelkd. 24: 49-54. BRAGG, D. B., H. A. SALEM, AND T. J. DEVLIN. BRUNE, G., E. THIER, AND E. BORCHERT. 1968. 1970. Effect of dietary triticale ergot on the per­ Variability of biological value of protein of different formance and survival of broiler chicks. Can. J. types of cereal (variants with manuring and site). Anim. Sci. 50: 259-264. 3. Metabolic trials with pigs and rats and comparison BRAGG, D. B., AND T. F. SHARBY. 1970. Nutritive with amino acid scores (in German). z. Tierphysiol. value of triticale for broiler chick diets. Poult. Sci. Tiererenaehr. Futtermittelkd. 24: 89-107. 49(4): 1022-1027. BRUNO, D., AND K. J. CARPENTER. 1957. A modi­ BRESsANI, R. 1971. Amino acid supplementation fied procedure for the estimation of ·available of cereal grain flours tested in children. pp. 184-204. lysine' in food proteins. Biochem. J. 67: 13P. In N. S. Scrimshaw and A. M. Altschul [ed.] Amino BURKE, R. P. 1960. The effect of bread baking and acid fortification of protein foods. Report of an drying on the stability of lysine, histidine and international conference. Massachusetts Institute of arginine. Nutr. Soc. (South Africa) Proc. I : 63-64. Technology, Cambridge, Massachusetts, USA. BUSCH, R. H., AND H. D. WILKINS. 1972. Agro­ ISBN 0 262 19091 5. nomic performance of triticale in North Dakota. BRESSANI, R., AND F. VITERI. 1970. Metabolic studies North Dakota Farm. Res. 29(3): 29-30. in human subjects. pp. 344-357. In Proc. SOS/70 BusHUK, W. 1972. Review of the literature on the Third Int. Cong. Food Sci. Technol. Washington, biochemical, technological and nutritional proper­ D.C., USA. ties of triticale. Background material for the 162 MONOGRAPH IDRC-02Ie

Triticale Utilization Work Group, International CHEN, C. H., AND W. BusHUK. 1970a. Nature of Development Research Centre. University of Mani­ proteins in Triticale and its parental species. I. toba, Winnipeg, Canada. Solubility characteristics and amino acid com­ BUSHUK, W., P. HWANG, AND C. W. WRIGLEY. 1971. position of endosperm proteins. Can. J. Plant Sci. Proteolytic activity of maturing wheat grain. Cereal 50(1): 9-14. Chem. 48: 637-639. l 970b. II. Gel filtration and disc electrophoresis CALHOUN, W. K., F. N. HEPBURN, AND W. B. BRADLEY. results. Can. J. Plant Sci. 50(1): 15-24. 1960. The availability of lysine in wheat, flour, l 970c. III. A comparison of their electrophoretic bread and gluten. J. Nutr. 70(3): 337-347. patterns. Can. J. Plant Sci. 50(1): 25-30. CALVEL, R. 1962. The Modern Bakery (in French). CHICK, H., M. E. M. CUTTING, C. J. MARTIN, AND 3rd ed. Eyrolles, Paris. 466 p. E. B. SLACK. 1947. Observations on the digesti­ CAMPBELL, J. A. 1963. Methodology of protein bility and nutritive value of the nitrogenous con­ evaluation. Publ. 21, American Univ. Beirut, stituents of wheat bran. Br. J. N utr. 1 : 161-182. Lebanon. CHOWDHARY, s. 1954. Indian Cookery. Andre CAMPBELL, J. A., AND D. G. CHAPMAN. 1959. Deutsch, London. Evaluation of proteins in food-criteria for des­ CIMMYT 1970-71. Report on Maize and Wheat cribing protein value. J. Can. Diet. Assoc. 21: 51-53, improvement. Annual Report. Centro Internacional 56-60. de Mejoramiento de Maiz y Trigo. Mexico 6. 114 p. CAMPBELL, J. A., AND J. M. MCLAUGHLAN. 1970. 1971. Bibliography of Wheat. [3 vol.) Inter­ Applicability of Animal Assays to Humans. pp. national Maize and Wheat Improvement Center. 336-343. In Proc. 3rd Int. Cong. Food Sci. Technol., Mexico. Scarecrow Press, Inc., Metuchen, N .J ., Washington, D.C. USA. ISBN 0-8108-0393-3. CARPENTER, K. J. 1960. The estimation of available CLARK, H. E., J. M. HOWE, E. T. MERTZ, AND L. L. lysine in animal protein foods. Biochem. J. 77: 604-- REITZ. 1959. Lysine in baking powder biscuits. 610. J. Am. Diet. Assoc. 35(5): 469-471. 1970. Nutritional considerations in attempts to CLARK, H. E., J. M. HOWE, B. M. SHANNON, K. change the chemical composition of crops. Proc. CARLSON, ANDS. M. KOLSKI. 1970. Requirements Nutr. Soc. 29 (I): 3-12. of adult human subjects for methionine and cystine. CARPENTER, K. J ., AND H.J. H. DE MUELENAERE. 1965. Am. J. Clin. Nutr. 23(6): 731-738. A comparative study of performance on high­ CLARKE, J-A. K., AND B. M. KENNEDY. 1962. protein diets of unbalanced amino acid composition. Availability of lysine in whole wheat bread and in Proc. Nutr. Soc. 24 (2): 202-209. selected breakfast cereals. J. Food Sci. 27: 609-616.­ CARPENTER, K. J., AND G. M. ELLINGER. 1955. The CLEGG, K. M. 1960. The availability of lysine in estimation of "available lysine" in protein con­ groundnut biscuits used in the treatment of kwashi­ centrates. Biochem. J. 61: XI-XII. orkor. Br. J. Nutr. 14: 325-329. CARPENTER, K. J., B. E. MARCH, C. K. MILNER, AND CLEGG, K. M., AND N. DAVIES. 1958. "Available R. c. CAMPBELL. 1963. A growth assay with lysine" in wheat, flour and bread. Proc. Nutr. Soc. chicks for the lysine content of protein concentrates. 17: x. Br. J. Nutr. 17: 309-323. COLE, E. R. 1969. Alternative methods to the CATE, R. B., AND L. A. NELSON. 1965. A rapid Kjeldahl estimation of protein nitrogen. Rev. Pure method for correlation of soil test analyses with Appl. Chem. 19: 109-130. plant response data. Inter. Soil Testing Series Tech. CONKERTON, E. J., AND V. L. FRAMPTON. 1959. Bull. I. Reaction of gossypol with free epsilon-amino groups CAVE, N. A. G., J. D. SUMMERS, S. J. SLINGER, AND of lysine in proteins. Arch. Biochem. Biophys. G. C. ASHTON. 1965. The nutritional value of wheat milling by-products for the growing chick. 81(1): 130-134. Cereal Chem. 42(6): 533-538. COONS, C. M. 1968. Selected references on cereal CHAMBERLAIN, N., T. H. COLLINS, G. A. H. ELTON, grains in protein nutrition. Human and experimental D. F. HOLLINGSWORTH, D. B. LISLE, AND P. R. animal studies of major and minor cereals 1910- PAYNE. 1966. Studies on the composition of 1966. ARS 61-5. United States Department of food. 2. Comparison of the nutrient content of Agriculture, Washington, D.C. 150 p. bread made conventionally and by the Chorleywood CORNEJO, S., J. POTOCNJAK, AND D. W. ROBINSON. Bread Process. Br. J. Nutr. 20: 747-755. 1972. Comparative nutritional value of triticale for CHAPMAN, D. G., R. CASTILLO, AND J. A. CAMPBELL. swine. J. Anim. Sci. 34(5): 893. 1959. Evaluation of protein in foods. I. A method COUCH, J. R., AND F. G. HOOPER. 1972. p. 183. for the determination of protein efficiency ratios. In A. A. Albanese[ed.]. Newer methods of nutritional Can. J. Biochem. Physiol. 37(5): 679-686. biochemistry. Academic Press. New York. REFERENCES 163

CREEK, R. D., AND v. VASAITIS. 1962. Detection of menting rice and wheat and diets based on them an antiproteolytic substance in raw wheat germ. with L-lysine and DL-threonine on the nutritive Poult. Sci. 41: 1351-1352. value of their proteins. J. Nutr. Diet. 5: 278-282. CREMER, H. D., K. LAND, AND K. HERGT. 1953. The DAWBARN, M. C. 1949. The effects of milling upon improvement in the nutritive value of bread (in the nutritive value of wheaten flour and bread. German). Brot Geback 7: 69. Nutr. Abstr. Rev. 18(4): 691-706. CROY, L. I., AND R. H. HAGEMAN. 1970. Relation­ DEDIO, W., P. J. KALTSIKES, AND E. N. LARTER. 1969. ship of nitrate reductase activity to grain protein A thin-layer chromatographic study of the phenolics production in wheat. Crop Sci. 10: 280--285. of triticale and its parental species. Can. J. Bot. CUBADDA, R., A. FRATONI, AND E. QUATTRUCCI. 47(10): 1589-1593. 1970. Determining the nutritive value of alimentary DE LA PLAZA, S., J. M. VALLEJO, P. CASASECA. AND pastes dried at various temperatures. I. Biological F. GARCIA OLMEDO. 1969. Utilization of hexa­ tests with fine flour pastes (in Italian). Quad. Nutr. ploid triticale. I. Experimental milling (in Spanish). 30(4-6): 134-143. An. Invest. Agron. 18(4): 357-370. CuuK, R., AND H. R. ROSENBERG. 1958. The forti­ DEMAN, J. M., E. 0. I. BANIGO, V. RASPER, H. GADE, fication of bread with lysine. 4. The nutritive value ANDS. J. SLINGER. 1973. Dehulling of sorghum of lysine-supplemented bread in reproduction and and millet with the Palyi compact milling system. lactation studies with rats. Food Tech. 12(4): J. Int. Can. Sci. Technol. Aliment. 6(3): 188-193. 169-174. DE MUELENAERE, H. J. H., M. L. CHEN, AND A. E. DALBY, G. 1969. Protein fortification of bread: HARPER. 1967. Assessment of factors influencing Baladi bread as an example. pp. 174-180. In Max estimation of availability of threonine, isoleucine Milner [ed.] Protein enriched cereal foods for world and valine in cereal products. J. Agric. Food Chem. needs. American Association of Cereal Chemists. 15: 318-323. St. Paul, Minnesota 55104, USA. DEODIKAR, G. B. 1963. Rye (Seca/e cerea/e Linn.). DALRYMPLE, D. G. 1972. Imports and plantings of Indian Council of Agric. Res. Cereal Crop Ser. No. high yielding varieties of wheat and rice in the less Ill. Indian Council of Agricultural Research, New developed nations. pp. 48-51. In FEDR-14. United Delhi, India. States Department of Agriculture. DEOSTHALE, Y. G., K. SURYANARAYANA RAO, AND DALRYMPLE, D. G., AND w. I. JONES. 1973. Evaluat­ V. S. MOHAN. 1969. Nutritive value of some ing the "green revolution." Paper presented at varieties of wheat. J. Nutr. Diet. 6: 182-186. Nutrition and New Food Technology session of the DE VUYST, A., W. VERVACK, M. VANBELLE, R. first meeting of the American Association for the ARNOULD, AND A. MOREELS. 1958. The amino Advancement of Science and the Consejo Nacional acid composition of Belgian cereals and their value de Ciencia y Technologia, Mexico City, June 20, in feeding pigs and poultry (in French). Agric. Louv. 1973. 6(1): 19-53. DANIEL, V. A., B. L. M. DESAI, S. VENKAT RAO, M. DHARAM JIT SINGH. 1956. Classic cooking from SwAMINATHAN,ANDH. A. B. PARPIA. 1969. Mutual India. Riverside Press. Cambridge, Mass., U.S.A. and amino acid supplementation of proteins. IV. The nutritive value of the proteins of blends of wheat DIFCO LABORATORIES INC. Difeo Manual 9th ed. Detroit, Mich. and Bengal gram fortified with limiting amino acids. J. Nutr. Diet. 6: 15-21. D1NCA, D., H. CIMPEANU, T. TABARANU, AND M. DANIEL, V. A., T. R. DORAISWAMY, S. VENKAT RAO, ARMEANU. 1971. The influence of rotation on M. SwAMINATHAN, AND H. A. B. PARPIA. 1968a. yield, utilization of fertilizers and the biological The effect of supplementing a poor wheat diet with quality of wheat and com crops on reddish-brown L-lysine and DL-threonine on the digestibility co­ forest soil (in Romanian). Probleme agric. 23(9): efficient, biological value and net utilization of 12-22. proteins and nitrogen retention in children. J. Nutr. DONOSO, G., M. MUNOZ, I. BARJA, E. DURAN, M. Diet. 5: 134-141. URREA, AND J. v. SANTA MARIA. 1963. Enrich­ DANIEL, V. A., R. LEELA, T. S. SUBRAMANYARAJ URS, ment of bread with fish flour for human consump­ S. VENKAT RAO, D. RAJALAKSHMI, AND M. SwAMl­ tion. I. Study of breadmaking and acceptability NATHAN. 1965. The supplementary value of the trials. Nutr. Bromatol. Toxicol. 2: 75-84. proteins of soya bean as compared with those of DuBETZ, S. 1972. Effects of nitrogen on yield and Bengal gram, red gram and skim milk powder to protein content of Manitou and Pitic wheats grown poor Indian diets based on rice and wheat. J. Nutr. under irrigation. Can. J. Plant Sci. 52: 887-890. Diet 2: 128-133. DUFFIELD, R. D., L. I. CROY, AND E. L. SMITH. 1972. DANIEL, V. A., S. VENKAT RAO, M. SwAMINATHAN, Inheritance of nitrate reductase activity, grain pro­ AND H. A. B. PARPIA. 1968b. Amino acid supple­ tein, and straw protein in a hard red winter wheat mentation of proteins. 111. The effect of supple- cross. Agronomy J. 64(2): 249-251. 164 MONOGRAPH IDRC-021e

EDWARDS, C. H., L. K. BOOKER, C. H. RUMPH, W. G. tion of Nuclear Techniques. International Atomic WRIGHT, AND S. N. GANAPATHY. 1971. Utiliza­ Energy Agency, Vienna. tion of wheat by adult man: nitrogen metabolism, FEILLET, P. 1965. Contribution to the study of the plasma amino acids and lipids. Am. 1. Clin. Nutr. proteins of wheat; influence of genetic, agronomic 24(2): 181-193. and technological factors (in French). Ann. Technol. EFREMOV, V. V., E. M. MASLENIKOVA, Yu. M. NEMEN­ Agric. 14 (numero hors ser. I): 1-94. OVA, L. G. GvozDOVA, E. A. KRAIKO, L. S. KRUMS, FELLERS, D. A., A. D. SHEPHERD, N. 1. BELLARD, AND 0. I. PENAR, A. SH. VAINERMAN. 1970. Biological A. P. MOSSMAN. 1966. Protein concentrates by value of wheat bread containing various food dry milling of wheat mill feeds. Cereal Chem. 43(6): adjuvants. Gig. i Sanitariya 35(6): 88-90. 715-725. EGGUM, B. 0. 1970. Current methods of nutritional FERHI, Y. 1970. Grandes Recettes de la Cuisine protein evaluation, pp. 289-302. In Proc. Joint Algerienne. S.N.E.D. Paris. IAEA/FAO Symp. on Plant Protein resources: FERNANDEZ, R., S. M. KIM, E. R1vERA, AND 1. Mc­ their improvement through the application of GINNIS. 1972. The use of rye and triticale as main nuclear techniques. International Atomic Energy ingredients in the diets of laying hens. Poult. Sci. Agency. Vienna. 51(5): 1806. 1973. The levels of blood amino acids and blood FERREL, R. E., A. D. SHEPHERD, AND D. G. GUADAGNI. urea as indicators of protein quality, pp. 317-327. 1970. Storage stability of lysine in lysine-fortified In 1. G. W. Porter and B. A. Rolls [ed.] Proteins in wheat. Cereal Chem. 47(1): 33-37. human nutrition. Academic Press, London. FINNEY, D. F., M. D. SHOGREN, Y. POMERANZ, AND EL GINDY, M. M., c. A. LAMB, AND R. c. BURRELL. L. C. BoL TE. 1971. Cereal malts in breadmaking. 1957. Influence of variety, fertilizer treatment and Bakers Dig. 46: 36--38, 55. soil on the protein content and mineral composition FISHER, L. 1. 1972. Evaluation of triticale silage for of wheat, flour and flour fractions. Cereal Chem. lactating cows. Can. 1. Anim. Sci. 52: 373-376. 34(3): 185-195. FLODIN, N. W. 1959. Lysine supplementation of EL-LAKANY, S., 1. BIELY, AND B. E. MARCH. 1969. wheat foods. Assoc. Food Drug Off. US Q. Bull. Improved nutrient utilization from wheat subjected 23(2): 76--89. to autoclaving and freezing. Cereal Chem. 46(3): FLOUR MILLING AND BAKING RESEARCH ASSOCIATION. 301-304. 1971. Annual Report. p. 14. Flour Milling and ELLIOTT, F. C. 1963. Quart. Bull. Michigan Agric. Baking Research Association. Chorleywood, Herts., Exp. Stn. 46: 58. England. ELLIS, R. P. 1971. Electrophoresis of grain proteins: FOOD AND AGRICULTURE ORGANIZATION. 1959. Food detection of rye proteins in wheat x rye hybrids. additive control in Canada. FAO Food Additive 1. Natl. Inst. Agric. Bot. 12: 236--241. Control Series I. Rome. 36 p. ELONEN, P., L. AHO, AND P. KOIVISTOINEN. 1972. 1960. Food additive control in the United King­ Influence of irrigation and nitrogen fertilization on dom. FAO Food Additive Control Series 2. Rome. the amino acid composition of spring wheat. 52 p. Maataloustiet. Aikak. 44(1): 56--62. 1961 a. Food additive control in the Netherlands. ERMOLAEv, I. 1970. Effect of the sowing norm, FAO Food Additive Control Series 3. Rome. 41 p. time of sowing and the rate of fertilization on certain 1961 b. Food additive control in Australia. FAO biological properties and the yield of Bezostaya I Food Additive Control. Series 4. Rome. 30 p. wheat variety (in Bulgarian). Raskenievodudri Nauk. 1961c. Food additive control in Denmark. FAO (Plant Sci.). 7(9): 83-96. Food Additive Control Series 5. Rome. 35 p. EWALD, E., AND G. WENZEL. 1967. Effect of nitro­ 1963a. Food additive control in France. FAO gen fertilization on protein composition and amino Food Additive Control Series 6. Rome. 69 p. acid content of wheat grain [in German]. Qual. 1963b. Food additive control in the Federal Plant. Mater. Veg. 14: 98-104. Republic of Germany. FAO Food Additive Control FANCE, W. 1. 1968. The international confectioner. Series 7. Rome. 41 p. Virtue & Co. Ltd., London. 1965. Protein requirements. FAO Nutr. Meet. FANCE, W. 1., AND B. H. WRAGG. 1968. Up-to-date Rep. 37. Food and Agriculture Organization, Rome. breadmaking. Maclaren and Sons Ltd., London. I 969a. Provisional indicative world plan for agri­ 297 p. cultural development. c 69/4(1) Food and Agri­ FAVRET, E. A., L. MANGHERS, R. SOLARI, A. AVILA, culture Organization, Rome. AND 1. c. MONESIGLIO. 1970. Gene control of 1969b. Food additive control in the' USSR. protein production in cereal seeds pp. 87-97. In FAO Food Additive Control Series 8. Rome. 45 p. Proc. Joint IAEA/FAO Symp. on Plant Protein 1970a. Production Yearbook Vol. 24. Food and Resources: their improvement through the applica- Agriculture Organization. Rome. 822 p. REFERENCES 165

I 970b. Amino acid content of foods and bio­ GONTZEA, I., AND P. SUTZESCU. 1968. Natural anti­ logical data on proteins. FAQ Nutr. Studies No. 24. nutritive substances in foodstuffs and forages. K. Food and Agriculture Organization, Rome. 285 p. Karger, Basel (Switzerland). I 970c. Composite flour programme. Develop­ GONTZEA, I., P. SUTZESCU, AND F. POPESO. 1970. ment of bakery products and paste goods from The effect of extraction rate on the nutritive value cereal and non-cereal flours, starches and protein of wheat flour and the possibility of its improvement concentrates. Documentation Package No. I. Food by the addition of defatted wheat germ (in French). and Agriculture Organization. Rome, Italy. 176 p. Vitals!. Zivilisationskr. 15(2): 63-66. 1971. Meeting Report on Single Cell Protein. GOTTHOLD, M. L. McG., AND B. M. KENNEDY. 1964. Document 3. 14/15. Moscow. Food and Agriculture Biological evaluation of protein in steamed and Organization. Rome. baked breads and in bread ingredients. J. Food Sci. FOOD AND DRUG LABORATORIES. 1966. Determina­ 29: 227-232. tion of protein rating. Official Method F0-40. GRACZA, R. 1959. The subsieve-size fractions of a Health Protection Branch, Health and Welfare soft wheat flour produced by air classification. Canada. Cereal Chem. 36: 465-487. FORD, J. E. 1962. A micro biological method for GRAHAM, G. G. 1971. Methionine or lysine forti­ assessing the nutritive value of proteins. 2. The fication of dietary protein for infants and small measurement of "available" methionine, leucine, children. pp. 222-236. /n N. S. Scrimshaw and A. M. isoleucine, arginine, histidine, trytophan and valine. Altschul [ed.] Amino acid fortification of protein Br. J. Nutr. 16: 409-425. foods. MIT Press, Cambridge, Massachusetts. ISBN 1964. A micro biological method for assessing 0 262 19091. the nutritive value of proteins. 3. Further studies on GRAHAM, G. G., A. CORDANO, AND J. M. BAERTL. the measurement of available amino acids. Br. J. 1966. Studies in infantile malnutrition IV. The Nutr. 18: 449-460. effect of protein and calorie intake in serum proteins. FRAENKEL-CONRAT, H., AND M. COOPER. 1944. The Am.J.Clin.Nutr.18: 11-15. use of dyes for the determination of acid and basic GRAHAM, G. G., A. CORDANO, E. MORALES, G. groups in proteins. J. Biol. Chem. 154(1): 239-246. ACEVEDO, AND R. P. PLACKO. 1970. Dietary FRASER, J. R., AND D. c. HOLMES. 1959. Proximate protein quality in infants and children. V. A wheat analysis of wheat flour carbohydrates. 4. Analysis of flour-wheat concentrate mixture. Plant Foods for wholemeal flour and some of its fractions. J. Sci. Human Nutr. 2(1): 23-27. Food Agric. 10: 506--512. GRAHAM, G. G., R. P. PLACKO, G. ACEVEDO, E. FRIEND, D. W. 1970. Comparison of some milling MORALES, AND A. CORDANO. 1969. Lysine en­ products of barley and rye when fed in diets to rats. richment of wheat flour: evaluation in infants. Can. J. Anim. Sci. 50(2): 345-348. Am. J. Clin. Nutr. 22(11): 1459-1468. GANDHI, S. M., AND K. S. NATHAWAT. 1968. GREER, E. N., AND G. G. GRINDLEY. 1954. The Influence of nitrogen fertilization on quality charac­ late manuring of winter wheat: an observation on ters in a few varieties of common wheat (Triticum the nutritive value of the grain. J. Agric. Sci. 45(1): aestirum L.). Indian J. Agric. Sci. 38(1): 47-52. 125-128. GATES, J.C., AND B. M. KENNEDY. 1964. Protein GREWE, E., AND J. A. LECLi;RC. 1943. Wheat, quality of bread and bread ingredients. J. Am. Diet. wheat germ in breadmaking. Cereal Chem. 20: 434-- Assoc. 44: 374--377. 447. GERMANN, A. F. 0., E.T. MERTZ, AND W. M. BEESON. GRISWOLD, R. M. 1951. Effect of heat on the 1958. Revaluation of L-lysine requirement of nutritive value of proteins. J. Am. Diet. Assoc. weanling pigs. J. Anim. Sci. 17: 52-61. 27(2): 85-93. GHAI, 0. P., ANDS. N. CHAUDHURI. 1971. Feeding GUENTHNER, E., AND c. w. CARLSON. 1970. A com­ trials with lysine fortified wheat flour in protein parison of triticale, corn, wheat and milo laying calorie malnutrition. Indian J. Med. Res. 59(5): diets. Poult. Sci. 49(5): 1390 (Abstr.). 756--759. GUGGENHEIM, K., AND N. FRIEDMANN. 1960. Effect GIRISH, G. K., S. K. SAHNI, S. K. GUPTA, AND K. of extraction rate of flour and of supplementation KRISHNAMURTHY. 1971. Analysis of wheat and with soya meal on the nutritive value of bread wheat products from roller flour mills in India. protein. Food Technol. Champaign. 14: 298-300. Bull. Grain Technol. 9(1): 26--35. GUGGENHEIM, K., E. PEREZ, J. ILAN, S. JosEPH, AND GOLENKOV, V. F., AND V. M. GILZIN. 1971. Study A. GOLDBERG. 1962. The nutritive value of pro­ of the amino acid composition of the protein of teins of human diets as affected by extraction rate winter rye grain (in Russian). Prikl. Biokhim. of flour and its supplementation with soya meal. Mikrobiol. 7(3): 328-333. Am. J. Clin. Nutr. 10(3): 231-239. 166 MONOGRAPH IDRC-021e

GUNDEL, J., M. A. REGIUS, G. M. SzELENYI, AND B. tional status of school children. Acta Paediat. TOTH. 1970. Studies on the nutritive value of Scand. 62: 297-303. triticale. I. Chemical and physiological studies (in HEGSTED, D. M. 1971. Nutritional research on the Hungarian). Allattenyesz. 19(2): 171-178. value of amino acid fortification -experimental GUNTHARDT, H., AND J. McGINNIS. 1957. Effect of studies in animals, pp. 157-178. In N. S. Scrimshaw nitrogen fertilization on amino acids in whole wheat. and A. M. Altschul [ed.]. Amino acid fortification J. Nutr. 61(2): 167-176. of protein foods. MIT Press, Cambridge, Masssa­ GUPTA, Y. P. 1970. Preliminary studies on lysine chusetts. and methionine content of dwarf and improved 1972. (in press). In Proc. Symp. on Protein in varieties of Indian wheats. Sci. Cult. 36( 12): 652-654. Processed Foods. Council on Foods and Nutrition, HAENEL, H., ANDS. G. KHARATYAN. 1973. Some American Medical Assoc., Chicago. observations on the use of microbiological tech­ HEGSTED, D. M., AND Y. CHANG. 1965a. Protein niques for the determination of protein quality, pp. utilization in growing rats. I. Relative growth in­ 195-205. In J. G. W. Porter and B. A. Rolls [ed.]. dex as a bioassay procedure. J. Nutr. 85: 159-168. Proteins in human nutrition. Academic Press, l 965b. Protein utilization in growing rats at London. different levels of intake. J. Nutr. 87: 19-25. HALE, H. E. 1963. Production aspects of protein HEGSTED, D. M., AND J. WORCESTER. 1966. Assess­ speciality breads. Bakers Dig. 37(3): 61-62, 65-66. ment of protein quality with young rats. Proc. HALEVY, S., AND N. GROSSOWICZ. 1953. A micro­ Seventh Int. Congress Nutr. biological approach to nutritional evaluation of HEGSTED, D. M., R. NEFF, AND J. WORCESTER. 1968. proteins. Proc. Soc. Exp. Biol. Med. 82: 567-571. Determination of the relative nutritive value of pro­ HARPER, A. E. 1969. Amino acid toxicities and teins. Factors affecting precision and validity. J. imbalances, pp. 391-422. In H. N. Munro [ed.]. Agric. Food Chem. 16(2): 190-195. Mammalian protein metabolism. Vol. 3. Academic HEGSTED, D. M., M. F. TRULSON, H. S. WHITE, P. L. Press, New York. WHITE, E. VINAS, E. ALVISTUR, C. DIAZ, J. VASQUEZ, HARPER, A. E., AND Q. R. ROGERS. 1965. Amino A. Loo, A. ROCA, CH. CoLLAZOS, AND A. RUIZ. acid imbalance. Proc. Nutr. Soc. 24(2): 173-190. 1955. Lysine and methionine supplementation of HARROLD, R. L., W. E. DINUSSON, C. N. HAUGSE, AND all-vegetable diets for human adults. J. Nutr. 56: M. L. BUCHANAN. 1971. Triticale as feed for 555-576. growing-finishing swine. North Dakota Farm Res. HENRY, K. M., ANDS. K. KoN. 1949. The effect on Bi-mon. Bull. 28(3)·. 34-36. the biological value of bread nitrogen of additions HAUNOLD, A., V. A. JoHNSON, AND J. W. SCHMIDT. of dried skim milk and of soya flour. J. Dairy Res. l 962a. Variation in protein content of the grain 16: 53-57. in four varieties of Triticum aestirium L. Agron. J. HEPBURN, F. N., AND W. B. BRADLEY. 1965. The 54: 121-125. amino acid composition of hard wheat varieties as l 962b. Genetic measurements of protein in the a function of nitrogen content. Cereal Chem. 42(2): grain of Triticum aestirium L. Agron. J. 54: 203-206. 140-149. HAY, J. G. 1942. The distribution ofphytic acid in HEPBURN, F. N., W. K. CALHOUN, AND W. B. BRADLEY. wheat and a preliminary study of some of the l 960a. A growth response of rats to glutamic acid calcium salts of this acid. Cereal Chem. 19(3): when fed an amino acid diet. J. Nutr. 72(2): 163-168. 326-333. l 960b. The distribution of the amino acids of HEDAYAT, H. 1971. Lysine field trial in Iran, pp. wheat in commercial mill products. Cereal Chem. 282-288. In N. S. Scrimshaw and A. M. Altschul 37(6): 749-755. [ed.]. Amino acid fortification of protein foods. 1966. The biological availability of essential MIT Press, Cambridge, Massachusetts. ISBN amino acids in wheat, flour, bread and gluten. 0262 190915. Cereal Chem. 43(3): 271-283. HEDAYAT, H., N. SARKISSIAN, S. LANKARANI, AND G. HEPBURN, F. N., E.W. LEWIS, JR., ANDC. A. ELVEHJEM. DoNoso. 1968. The enrichment of whole wheat 1957. The amino acid content of wheat, flour and flour and Iranian bread with lysine and vitamins. bread. Cereal Chem. 34(5): 312-322. Acta Biochim. Iranica. 5: 16-25. HILL, F. W., AND D. L. ANDERSON. 1958. Com­ HEDAYAT, H., H. SHAHBAZ!, R. PAYAN, AND G. parison of metabolizable energy and productive DoNoso. 1971. Lysine field trial in Iran, pp. energy determination with chickens. J. Nutr. 64: 44-52. In S. Margen [ed.]. Progress in Human 587-604. Nutrition. Vol. I. A VI Pub!. Co., Westport, Conn. HILL, F. W., D. L. ANDERSON, R. RENNER, AND L. B. HEDAYAT, H., H. SHAHBAZ!, R. PAYAN, M. AZAR, M. CAREW, JR. 1960. Studies of metabolizable energy BAVENDI, AND G. DONOSO. 1973. The effect of of grain and grain products for chickens. Poult. lysine fortification of Iranian bread on the nutri- Sci. 39: 573-579. REFERENCES 167

HINTON. J. J.C. 1953. The distribution of protein HULSE, J. H. (in press). Protein supplementation of in the maize kernel in comparison with that in wheat. bread and baked foods. In A.A. Altschul [ed.]. New Cereal Chem. 30(5): 441-445. Protein Foods. Academic Press Inc .. 111 Fifth HOFFMAN, W. s .. AND G. c. McNEIL. 1949. The Avenue. New York, N.Y. 10003. enhancement of the nutritive value of wheat gluten HUMMEL, C. H. 1966. Macaroni Production. Food by supplementation with lysine, as determined from Trade Press, London. nitrogen balance indices in human subjects. J. Nuir. HUNDLEY, J.M., R. B. ING, AND R. W. KRAUSS. 1956. 38: 331-343. Algae as sources of lysine and threonine in supple­ HOJJATI, S. M .. AND M. MALEK!. 1972. Effect of menting wheat and bread diets. Science 124: 536-- potassium and nitrogen fertilization on lysine, 537. methionine and total protein contents of wheat HUTCHEON, W. L., AND E. A. PAUL. 1966. Control grain, Triticum aestirnm L. em Thell. Agron. J. of the protein content of Thatcher wheat by nitrogen 64( I): 46--48. fertilization and moisture stress. Can. J. Soil Sci. HORAN, F. E. 1967. Defatted and full-fat soy flours 46(2): 101-108. by conventional processes. US Agric. Res. Ser. HUTCHINSON, J. B., AND H. F. MARTIN. 1970. ARS-71-35: 129-141. May 1967. Nutritive value of wheat bran. I. Effects of fine HORN, M. J., A. E. BLUM, C. E. F. GERSDORFF. AND grinding upon bran, and of added bran upon the H. W. WARREN. 1953. Sources of error in micro­ protein quality of white flour. J. Sci. Food Agric. biological determination of amino acids on acid 21(3): 148-151. hydrolysates. I. Effect of humin on amino acid HUTCHINSON, J. B., T. MORAN, AND J. PACE. 1956. values. J. Biol. Chem. 203: 907-913. Nutritive value of the protein of white and whole­ HORN, M. J., AND D. BREESE JONES. 1945. A rapid meal bread in relation to the growth of rats. Royal colorimetric method for the determination of trypto­ Soc. (London) Proc. [BJ 145: 270--279. phane in proteins and foods. J. Biol. Chem. 157: 1958. Bread and the growth of weanling rats: 153-160. the lysine-threonine balance. Nature, Lond. 181 : HORN, M. J., C. C. FIFIELD, A. E. BLUM, AND H. W. 1733-1734. WARREN. 1958. The distribution of amino acids 1959. The nutritive value of bread protein as in wheat and certain wheat products. Cereal Chem. influenced by the level of protein intakes, the level 35(6): 411-421. of supplementation with L-lysine and L-threonine, HOWARD, A. N., AND T. B. ANDERSON. 1965. The and the addition of egg and milk protein. Br. J. treatment of obesity, and bread of high protein and Nutr. 13(2): 151-163. low carbohydrate content. Proc. Nutr. Soc. 24(2): 1960. The quality of the protein in germ and XXVlll-XXIX. milk breads as shown by the growth of weanling 1968. The treatment of obesity with a high rats: the significance of the lysine content. J. Sci. protein loaf. Practitioner 201: 491-496. Food Agric. 11 (I 0): 576--582. HOWARD, H. W., W. J. MONSON, C. D. BAUER. AND 1963. The supplementation of wheat protein: R. J. BLOCK. 1958. The nutritive value of bread some general considerations. Milling 140(8): 194, flour proteins as affected by practical supplementa­ 197. tion with lactalbumin, nonfat dry milk solids, soy­ 1964. The effect of a steam treatment on the bean proteins, wheat gluten and lysine. J. Nutr. 64: feeding value of wheat. J. Sci. Food Agric. 15(6): 151-165. 413-417. HOWE, E. E., E. W. GILFILLAN, AND M. MILNER. INGALLS, J. R., T. J. DEVLIN, AND J. A. McKIRDY. l 965a. Amino acid supplementation of protein 1970. Triticale in diets for young dairy calves. concentrates as related to the world protein supply. Am. J. Clin. Nutr. 16: 321-326. Can. J. Anim. Sci. 50: 199-204. HOWE, E. E .• G. R. JANSEN, AND E. W. GILFILLAN. I NG ALLS, J. R .• AND G. D. PHILLIPS. 1971 . Effect of l 965b. Amino acid supplementation of cereal triticale ergot on calf growth. Can. J. Anim. Sci. grains as related to the world food supply. Am. J. 51(3): 819. Clin. Nutr. 16: 315-320. INSKEEP, G. C., A. J. WILEY, J. M. HOLDERBY, AND HRISTOVA, J.. AND R. BA EVA. 1972. Investigations L. P. HUGHES. 1951. Food yeast from sulfite on the nature of the protein phenotypes in triticale. liquor. Ing. Eng. Chem. 43(8): 1702-1711. Z. Pflanzenzuecht. 67: 327-336. IRVINE, G. N. 1971. Durum wheat and paste HUGHES, B. P. 1955. The intakes of essential amino products. pp. 777-796. In Y. Pomeranz [ed.]. Wheat acids of children who were deriving most of their Chemistry and Technology. Monograph Series, Vol. protein from bread and vegetables. Br. J. Nutr. I I I-revised. American Association of Cereal Chem­ 9: 373-378. ists, Inc., St. Paul, Minnesota. 168 MONOGRAPH IDRC-021e

IRWIN, M. I., AND D. M. HEGSTED. 1971. A con­ JOHNSON, V. A., J. W. SCHMIDT, AND P. J. MATTERN. spectus of research on amino acid requirements of I 968a. Cereal breeding for better protein impact. man. J. Nutr. 101(4): 539-566. Econ. Bot. 22: 16-25. JAHNKE, J. K., AND c. SCHUCK. 1957. Growth JOHNSON, V. A., J. W. SCHMIDT, P. J. MATTERN, AND response and liver fat deposition in rats fed bread A. HAUNOLD. 1963. Agronomic and quality

mixtures with varying levels of non-fat milk solids characteristics of high protein F 2-derived families and lysine. J. Nutr. 61(2): 307-316. from a soft red winter-hard red winter wheat cross. JAMALIAN, J., AND P. L. PELLETT. 1968. Nutritional Crop. Sci. 3(1): 7-10. value of Middle Eastern foodstuffs. IV. Amino acid JOHNSON, V. A., D. A. WHITED, P. J. MATTERN, AND composition. J. Sci. Food Agric. 19(7): 378-382. J. W. SCHMIDT. 1968b. Nutritional improvement JANICKI, J., AND J. KOWALCZYK. 1965. The bio­ of wheat by breeding. pp. 457-461. In Proc. 3rd Int. logical value of rye proteins compared with wheat Wheat. Genet. Symp. protein (in German). Vitals!. Zivilisationskr. 10: JONES, C.R., AND P. HALTON. 1958. High and low 14-21. protein flour. Their mechanics and utilization. JANICKI, J., J. WARCHALEWSKI, B. STASINSKA, AND J. Milling, 130: 100. KOWALCZYK. 1972. Amino acid compositions of JONES, C.R., P. HALTON, AND D. J. STEVENS. 1959. some flours (in Polish). Rocz. Technol. Chem. Zywn. The separation of flour into fractions of different 22(1): 71-78. protein contents by means of air classification. J. Biochem. Microbiol. Technol. Eng. 1(1): 77-97. JANSEN, G. R. 1969. Total protein value of protein JONES, D. B., A. CALDWELL, AND K. D. WIDNESS. and amino acid-supplemented bread. Am. J. Clin. 1948. Comparative growth promoting value of the Nutr. 22(1): 38-43. proteins of cereal grains. J. Nutr. 35(6): 639--{i50. JANSEN, G., S. R. EHLE, AND N. L. HAUSE. I 964a. JORDAN, R. M., AND H. E. HANKE. 1972. Finishing Studies of the nutritive loss of supplemental lysine lambs with triticale, barley or corn. Feedstuffs. in baking. I. Loss in standard white bread containing 44(19): 30. 4% non-fat dry milk. Food Technol. 18(3): 367-371. KALMYKOv, P. E. 1968. A comparative appraisal of I 964b. II. Loss in water bread and in breads rye and wheat proteins (in Russian). Vopr. Pitan. supplemented with moderate amounts of non-fat 27( I): 42-46. dry milk. Food Technol. 18(3): 372-375. KAO, C., AND R. J. ROBINSON. 1972. Aspergillus JANSEN, G., C. F. HUTCHISON, AND M. E. ZANETTI. flavus deterioration of grain: its effect on amino 1966. Supplementation of bread with fish flour acids and vitamins in whole wheat. J. Food Sci. and amino acids-a comparison of evaluation 37(2): 261-263. methods. Food Technol. 20(3): 91-94. KASARDA, D. D., C. C. NIMMO, AND G. 0. KOHLER. JOHNSON, D. W., AND L. S. PALMER. 1934. The 1971. Proteins and the amino acid composition of nutritive properties of protein, vitamins B and G, wheat fractions. pp. 227-299. In J. Pomeranz [ed.]. and the germ in rye. J. Agric. Res. 49(2): 169-181. Wheat chemistry and technology. American Associa­ JOHNSON, J. H. 1853. Preparation and application tion of Cereal Chemists, St. Paul, Minnesota, USA. of gluten. Brit. Patent Specif. No. 2004. KAUL, A. K., R. D. DHAR, AND P. RAGHAVIAH. 1970. JOHNSON, v. A., AND P. J. MATTERN. 1972. Sum­ The macro and micro dye-binding techniques of mary report of research findings from improvement estimating the protein quality in food samples. of the nutritional quality of wheat through increased J. Food Sci. Technol. 7(1): 11-16. protein content and improved amino acid balance. KENNEDY, B. M., AND R. SABISTON. 1960. Quality July I, 1966-Dec. 31, 1972. Agency Int. Dev., of the protein in selected wheat products. Cereal Dept. State, Washington, D.C., USA. Chem. 37( 4): 535-543. JOHNSON, V. A., P. J. MATTERN, AND J. W. SCHMIDT. KENT, N. L. 1966. Technology of cereals with 1967. Nitrogen relations during spring growth in special reference to wheat. Pergamon Press, Oxford, varieties of Triticum aestivium L. differing in the England. 262 p. protein content of their grain. Crop Sci. 7: 664-667. KENT-JONES, D. W., AND A. J. AMOS. 1967. Modern 1970. The breeding of wheat and maize with Cereal Chemistry. 6th ed. Food Trade Press Ltd., improved nutritional value. Proc. Nutr. Soc. 29(15): London. 730 p. 20--31. KHERA, M. S., H. C. KAPOOR, AND Y. P. GUPTA. JOHNSON, V. A., P. J. MATTERN, D. A. WHITED, AND 1972. Critical available soil phosphorus level of J. W. SCHMIDT. 1969. Breeding for high protein Indian soils in relation to protein quality of wheat. content and quality in wheat. pp. 29-40. In Proceed­ Agron. J. 64: 406-407. ings of a panel meeting on new approaches to KIES, C., AND H. M. Fox. I 970a. Effect of level of breeding for plant protein improvement. Inter­ total nitrogen intake on second limiting amino acid national Atomic Energy Agency, Vienna. in corn for humans. J. Nutr. 100: 1275-1285. REFERENCES 169

l 970b. Determination of the first limiting amino South Africa. A. A. Balkema, Cape Town, South acid of wheat and triticale grain for humans. Cereal Africa. Chem. 47(5): 615-625. KOFRANYI, E., AND H. MULLER-WECKER. 1960. Esti­ l 970c. Protein nutritive value of wheat and mation of the biological value of food proteins. IV. triticale grain for humans, studies at two levels of Comparison of the values of the proteins of milk, protein intake. Cereal Chem. 47(6): 671-678. rye and wheat with whole egg and calculations from 1972. Interrelationships of leucine with lysine, their amino acid composition (in German). Hoppe­ tryptophan, and niacin as they influence protein Seyler's Z. Physiol. Chem. 320: 233-240. value of cereal grains for humans. Cereal Chem. KOHLER, G. 0. 1964. Effect of processing on the 49(2): 223-231. nutritive value of wheat proteins. Cereal Sci. KIES, C., H. M. Fox, AND S. C-S. CHEN. 1972. Today 9: 173-178. Effect of quantitative variation in non-specific KOHLER, G. 0., AND R. PALTER. 1967. Studies on nitrogen supplementation of corn, wheat, rice and methods for amino acid analysis of wheat products. milk diets for adult men. Cereal Chem. 49( I): 26--33. Cereal Chem. 44(5): 512-520. KIES, C., H. M. Fox, AND E. R. WILLIAMS. 1967. KOHLER, G. 0., R. M. SAUNDERS, AND H. G. WALKER. Time, stress, quality and quantity as factors in the 1969. Digestibility of aleurone in wheat milling nonspecific nitrogen supplementation of com protein fractions. Cereal Sci. Today 14(3): Abstr. 88. for adult men. J. Nutr. 93: 337-385. KON, S. K., AND Z. MARKUZE. 1931. The biological KIES, C., E. WILLIAMS, AND H. M. Fox. 1965. values of the proteins of breads baked from rye and Effect of"non-specific" nitrogen intake on adequacy wheat flours alone or combined with yeasts or soya of cereal proteins for nitrogen retention in human bean flour. Biochem. J. 25: 1476--1484. adults. J. Nutr. 86: 357-361. KUIKEN, K. A., AND C. M. LYMAN. Availability of KIGER, J. L., AND J. G. KIGER. 1968. Modern amino acids in some foods. J. Nutr. 36(3): 359-368. techniques of the biscuit, cake and bread industry KURELEC, V., J. REGIUS, AND G. JECSAI. 1968. and craft, and of dietary products (in French). Nutritive value of feed wheats recently used in Vol. II. Dubod, Paris. 595 p. Hungary (in Hungarian). Allattenyesz. 17(3): KIHLBERG, R., AND L.-E. ERICSON. 1964. Amino 271-278. acid composition of rye flour and the influence of KURZEPA, H., J. BARTNIK, I. TRZEBSKA-JESKE, AND amino acid supplementation of rye flour and bread W. MORKOWSKA. 1960. Nutritive value of rye on growth, nitrogen efficiency ratio and liver fat in and its products. II (in Polish). Rocz. Panstw. Zaki. the growing rat. J. Nutr. 82: 385-394. Hig. 11(2): 81-101. KING, K. W., W. H. SEBRELL, JR., E. L. SEvERINGHAUS, LAFONT, P., AND J. LAFONT. 1970. Contamination AND W. 0. STORVICK. 1963. Lysine fortification of cereal products and animal feeds by aflatoxin (in of wheat bread fed to Haitian school children. Am. French). Food Cosme!. Toxicol. 8: 403-408. J. Clin. Nutr. 12(1): 36--48. LAKIN, A. L. l 973a. Evaluation of protein quality KLINGMAN, D. L. 1953. Effects of varying rates of by dye-binding procedures. pp. 179-193. /nJ. G. W. 2, 4-D and 2, 4, 5-T at different stages of growth on Porter and B. A. Rolls [ed.]. Proteins in human winter wheat. Agron J. 45: 606--610. nutrition. Academic Press, London. KNIGHT, J. W. 1965. Chemistry of wheat starch and l 973b. The estimation of protein and the evalu­ gluten and their conversion products. Leonard ation of protein quality by dye-binding procedures. Hill, London. Proc. Inst. Food Sci. Technol. (UK) 6(2): 80--83. KNIPFEL, J. E. 1969. Comparative protein quality LANG, L. 1950. The protein question and protein of triticale, wheat and rye. Cereal Chem. 46(3): enrichment of bread (in German). Getreide Mehl 313-317. Brot. 4(11/12): 120--122. KNIPFEL J. E., 1. R. AITKEN, D. C. HILL, B. E. LAOPIROJANA, P., S. ROBERTS, AND M. D. DAWSON. McDONALD, AND B. D. OwEN. 1971. Amino 1972. Nitrogen nutrition and yield relations of acid composition of food proteins: inter and intra­ Nugaines winter wheat. Agron. J. 64(5): 570--573. laboratory variation. J. Assoc. Off. Anal. Chem. LAPORTE, J., AND J. TREMOLIERES. 1962. Inhibitor 54(4): 777-781. action of rice, oats, maize, barley, wheat, rye and KOFRANYI, E. 1957. Estimation of the biological buckwheat flours on certain proteolytic enzymes of value of food proteins. Ill. Comparison of the the pancreas (in French). C. R. Soc. Biol. 156: biological value of milk protein with rye protein 1261-1263. (in German). Hoppe-Seyler's Z. Physiol. Chem. LARSEN, I., AND J. D. NIELSEN. 1966. The effect of 309: 253-265. varying nitrogen supplies on the content of amino 1971. The biological value of protein mixtures. acids in wheat grain. Plant Soil. 24(2): 299-307. pp. 345-353. In J. W. Claassens, and H.J. Potgieter, LARTER, E., T. TSUCHIYA, AND L. Ev ANS. 1968. [ed.]. Proteins and Food Supply in the Republic of Breeding and cytology of Triticale. pp. 213-221. In 170 MONOGRAPH IDRC-021e

K. W. Finlay and K. W. Shepherd [ed.]. Third Int. MCCOLLUM, E. V. 1945. Discussion [Bread "enrich­ Wheat Genet. Symp., Australia. ment"] Science 102: 181-182. LAWRENCE, J.M., K. M. DAY, E. HUEY, AND B. LEE. McDERMOTT, E. E., AND J. PACE. 1957. The content 1958. Lysine content of wheat varieties, species of amino acids in white flour and bread. Br. J. Nutr. and related genera. Cereal Chem. 35(3): 169-178. 11(4): 446-452. LEARMONTH, E. M., AND J. C. WOOD. 1963. A 1960. Comparison of the amino acid composi­ trypsin inhibitor in wheat. flour. Cereal Chem. tion of the protein in flour and endosperm from 40(1): 61-65. different types of wheat with particular reference to LIENER, I. E. 1969. Miscellaneous toxic factors. variation in lysine content. J. Sci. Food Agric. 11(2) pp. 409-448. In I. E. Liener [ed.]. Toxic constituents 109-115. of plant foodstuffs. Academic Press, New York. McDONALD, C. E., AND L. L. CHEN. 1964. Proper­ LIENER, I. E., AND M. KAKADE. 1969. Protease ties of wheat-flour proteinases. Cereal Chem. 4: inhibitors. pp. 8-68. In I. E. Liener [ed.]. Toxic 443-455. constituents of plant foodstuffs. Academic Press, MCELROY, L. W. 1968. Urea, biuret, triticale and New York. stilbestrol implants for finishing steers. Univ. LIGHT, R. F., AND c. N. FREY. 1943. The nutritive Alberta. 47th Annu. Animal Sci. Feeders Day Rep. value of white and whole wheat breads. Cereal Chem. 28. 20(6): 645-660. MCELROY, L. W., D. R. CLANDININ, W. LOBAY, AND LINDNER, K. 1956. Polarographic estimation of the S. I. PETHYBRIDGE. 1949. Nine essential amino biological value of protein (in German). Acta Chim. acids in pure varieties of wheat, barley, and oats. Acad. Sci. Hung. 9: 353. J. Nutr. 37: 329-336. 1964. Amino acid composition and nutritive McGINNIS, J. 1972. Biological evaluation of cereal value of wheat protein fractions (in German). grains for nutritional value: triticales, rye, wheat. Qua!. Plant. Mater. Veg. 11(1): 31-38. Rep. submitted to Rockefeller Foundation and the LOCKWOOD, J. F. 1960. Flour milling, 4th ed. Centro Internacional de Mejoramiento de Maiz Henry Simon Ltd., Stockport, Cheshire, England. y Trigo. Dept. Animal Sci., Washington State 526 p. University, Pullman, Washington 99163. LOFGREEN, G. P. 1969. Triticale, milo and barley MCLAUCHLAN, J. M., AND J. A. CAMPBELL. 1969. comparison (Progress report). Calif. Agric. Exp. Methodology of protein evaluation. pp. 391-422. Sta. Feeders Day Rep. p. 67. In H. N. Munro [ed.]. Mammalian protein metabo­ lism. Vol. 3. Academic Press, New York. LOMBARD, J. H., AND D. J. DE LANGE. 1965. The 1972. In press. In Proc. Symp. on Protein in chemical determination of tryptophan in foods and Processed Foods. Council on Foods and Nutrition, mixed diets. Anal. Biochem. 10: 260-265 American Medical Association, Chicago. LONNQUIST, J. H. 1969. High lysine maize. Protein MCLAUGHLAN, J.M., AND A. B. MORRISON. 1960. Advisory Group U.N. Document 2.31/3. Evaluation of protein quality in mixed foods. Can. LORENZ, K. 1972. Food uses of triticale, Food J. Biochem. Physiol. 38(11): 1378-1380. Technol. 26(11): 66, 68, 70, 72, 74. MCLAUGHLAN, J.M., C. G. ROGERS, D. G. CHAPMAN, LORENZ, K., AND J. MAGA. 1972. Triticale and AND J. A. CAMPBELL. 1959. Evaluation of protein wheat flour studies: composition of fatty acids, in foods. IV. A simplified chemical score. Can. J. carbonyls and hydrocarbons. J. Agric. Food Chem. Biochem. Physiol. 3 7(1): 1293-1299. 20(4): 769-772. MCNEAL, F. H., M. A. BERG, C. F. McGUIRE, V. R. LORENZ, K., W. DILSAVER, AND J. LOUGH. 1972a. STEWART, AND D. E. BALDRIDGE. 1972. Grain Evaluation of triticale for the manufacture of and plant nitrogen relationships in eight spring wheat noodles. J. Food Sci. 37(5): 764--767. crosses, Triticum aestirum L. Crop Sci. 12: 599-602. LORENZ, K., J. WELSH, R. NORMANN, AND J. MAGA. MADL, R. L., AND c. c. TSEN. 1973. Proteolytic l 972b. Comparative mixing and baking properties activity oftriticale. Cereal Chem. 50(2): 215-219. of wheat and triticale flours. Cereal Chem. 49(2): MALEK!, M., AND A. DJAZAYERI. 1968. Effect of 187-193. baking and arn'no acid supplementation on the MACAULIFFE, T., AND J. McGINNIS. 1971. Effect protein quality of Arabic bread. J. Sci. Food Agric. of antibiotic supplements to diets containing rye 19(8): 449-451. chick growth. Poult. Sci. 50(4): 1130-1134. MAMCHENKOV, I. P., AND G. V. PLATONOV. 1971. MCCANCE, R. A., AND E. M. WIDDOWSON. 1955. Importance of fertilizers in raising grain quality (in Breads White and Brown. J. B. Lippincott Co., Russian) Agrokhim. (7): 60-65. Philadelphia. 174 p. MATTERN, P. J., A. SALEM, v. A. JOHNSON, AND J. W. MCCLOY, A. W., L. B. SHERROD, R. C. ALBIN, AND SCHMIDT. 1968. Amino acid composition of K. R. HANSEN. 1971. Nutritive value of triticale selected high-protein wheats. Cereal Chem. 45(5): for ruminants. J. Anim. Sci. 32(3): 534--539. 437-444. REFERENCES 171

MATTERN, P. J., J. W. SCHMIDT, AND V. A. JoHNSON. Branch Canadian Society of Animal Produce 21 1970. Screening for high lysine content in wheat. (abstr.). Cereal Sci. Today 15(12): 409-411. MITCHELL, H. H. 1923-24. A method of determin­ MATTHEWS, R. H., G. RICHARDSON, AND H. LICHTEN­ ing the biological value of protein. J. Biol. Chem. STEIN. 1969 Effect of lysine fortification on 58(3): 873-903. quality of chapatties and yeast bread. Cereal Chem. 1925. The nutritive value of the proteins. Physiol. 46(1): 14-21. Rev. 4: 424-478. MATZ, S. A. 1968. Cookie and cracker technology. MITCHELL, H. H., AND R. J. BLOCK. 1946. Some Avi Publishing Co., Westport, Connecticut. 320 p. relationships between the amino acid contents of 1972. Bakery technology and engineering. 2nd proteins and their nutritive values for the rat. ed. Avi Publishing Co., Westport, Connecticut. J. Biol. Chem. 163(3): 599-620. 598 p. MITCHELL, H. H., AND G. C. CARMAN. 1926. The MAURON, J. 1961. The concept of amino acid biological value of the nitrogen of mixtures of patent availability and its bearing on protein evaluation. white flour and animal foods. J. Biol. Chem. 68: pp. 425-441. In Meeting protein needs of infants 183-215. and children. National Academy of Sciences­ MITCHELL, H. H., AND T. S. HAMILTON. 1929. The National Research Council. Washington, D.C. Biochemistry of the Amino Acids. American MAURON, J., AND F. MoTTu. 1962. Sweetened con­ Chemical Society Monograph Series, Chemical densed versus evaporated milk in improving the Catalog Co. Inc., New York. protein efficiency of wheat flour. J. Agric. Food MITSUDA, H. 1969. New approaches to amino acid Chem. 10(6): 512-515. and vitamin enrichment in Japan. pp. 208-219. In MAYR, R. 1948. The present application of soya Max Milner [ed.]. Protein-enriched cereal foods for products in German nutrition economy (in German). world needs. American Association of Cereal Getreide Mehl Brot (Sept): 140--141. Chemists, St. Paul, Minnesota, 55104. MEHDI, V. 1972. Amino acid composition of Indian MIZRAHI, S., G. ZIMMERMAN, Z. BERK, AND U. COGAN. wheats. Bull. Grain Technol. 10(1): 51-54. 1967. The use of isolated soybean proteins in MERTZ, E. T. 1971. Potential for genetic improve­ bread. Cereal Chem. 44(2): 193-203. ment of plant protein quality. pp. 96--102. In N. S. MOODY, E.G. 1973. Triticale in dairy cattle rations. Scrimshaw and A. A. Altschul [ed.]. Amino acid Feedstuffs 45(8): 38. fortification of protein foods. MIT Press, Cambridge, MOORE, S., AND W. H. STEIN. 1951. Chroma­ Massachusetts. tography of amino acids on sulfonated polystyrene MERTZ, E.T., L. S. BATES, AND 0. E. NELSON. 1964. resins. J. Biol. Chem. 192: 663-681. Mutant gene that changes protein and increases I 954a. Procedures for the chromatographic lysine content of maize endosperm. determination of amino acids in 4 per cent cross­ METTA, V. C. 1960. Nutritional value of fish flour linked sulfonated polystyrene resins. J. Biol. Chem. supplements. J. Am. Diet. Assoc. 37: 234-240. 211: 892-906. MILADI, S., D. M. HEGSTED, R. M. SAUNDERS, AND I 954b. A modified ninhydrin reagent for the G. 0. KOHLER. 1972. The relative nutritive value, photometric determination of amino acids and amino acid content and digestibility of the proteins related compounds. J. Biol. Chem. 211: 907-913. of wheat mill fractions. Cereal Chem. 49( I): 119-127. MOORE, S., D. H. SPACKMAN, AND W. M. STEIN. 1958. MILLER, D. S., AND A. E. BENDER. 1955. The Chromatography of amino acids on sulphonated determination of the net utilization of proteins by a polystryene resins. An improved system. Anal. Chem. shortened method. Br. J. Nutr. 9: 382-388. 30: 1185-1190. MILLER, D.S., AND P.R. PAYNE. 1961. Problems in the prediction of protein values of diets. The in­ MORAN, T. 1959. Nutritional significance of recent fluence of protein concentration. Br. J. Nutr. 15: work on wheat, flour and bread. Nutr. Abstr. Rev. 11-19. 29(1): 1-16. MILNER, c. K., AND K. J. CARPENTER. 1969. Effect MORRISON, A. B., AND J. A. CAMPBELL. 1960. of wet heat-processing on the nutritive value of whole Studies on th.: nutritional value of defatted fish wheat protein. Cereal Chem. 46(5): 425-434. flour. Can. J. Biochem. Physiol. 38: 467-473. MILNER, MAX. [ed.]. 1969. Protein-enriched cereal MossBERG, R. 1970. Practical problems concerning foods for world needs. American Association of the marketing of cereals with improved protein Cereal Chemists, St. Paul, Minnesota 55104. 343 p. value. Proc. Nutr. Soc. 29: 39-48. MINJA, L.A., D. A. CHRISTENSEN, AND F. W. SosULSKI. MuNCK, L. 1969. Genotype environment interac­ 1969. Nutritive value of several varieties and strains tion in protein production and utilization. pp. 173- of rye. Proc. Annu. Meeting General Society Eastern 184. In Proc. panel meeting on new approaches to 172 MONOGRAPH IDRC-02le

breeding for plant protein improvement. Inter­ A. A. Altschul [ed.). Amino acid fortification of national Atomic Energy Agency, Vienna. foods. MIT Press, Cambridge, Massachusetts. 1972. Improvement ofnutritional value in cereals. OLSEN, E. M. 1967. Effect of heat treatment on the Hereditas. 72: 1--128. quality and utilization by the rat of protein in wheat MuNCK, L., K-E. KARLSSON, AND A. HAGBERG. germ meal. Can. J. Biochem. 45: 1673-1679. 1969. pp. 544-558. In Second International Barley ORR, M. L., AND B. K. WATT. 1957. Amino acid Genetics Symposium. Washington State University, content of foods. Home Econ. Res. Rep. No. 4. Pullman, Washington. United States Department of Agriculture, Washing­ MuRAMATSU, K., H. TAKEUCHI, Y. FuNAKI, AND A. ton, D.C. CmsuwA. 1972. Influence of protein source on OSBORNE, T. B. 1924. The vegetable proteins. growth-depressing effect of excess amino acids Longmans Green, New York. in young rats. Agric. Biol. Chem. 36(8) 1269-1276. OSBORNE, T. B., L.B. MENDEL, AND E. L. FERRY. 1919. NAGY, D., W. WEIDLEIN, AND R. M. HIXON. 1941. A method of expressing numerically the growth­ Factors affecting the solubility of corn proteins. promoting value of proteins. J. Biol. Chem. 37(2): Cereal Chem. 18: 514--523. 223-229. NAIK, M. S. 1968. Lysine and tryptophan in pro­ OSER, B. L. 1951. Method for integrating essential tein fractions of sorghum. Indian J. Genet. Plant amino acid content in the nutritional evaluation of Breeding. 28(2): 142-146. protein. J. Am. Diet. Assoc. 27(5): 396-402. NAKAGAWA, I., T. TAKAHASI, T. SUZUKI, AND K. PAHWA, M. R., M. N. SADAPHAL, AND M. S. NAIK. KOBAYASHI. 1962. Amino acid requirements of 1972. Effect of crop rotations on the nutritive children: minimal needs of threonine, valine and quality of Sonora 64 wheat (Triticum aestil'ium L.). phenylalanine based on nitrogen balance method. Indian J. Agric. Sci. 42(7): 574--577. J. Nutr. 77: 61-68. PALMER, I. 1972. Food and the New Agricultural NARAYANASWAMY, D., S. KURIEN, V. A. DANIEL, S. Technology. Report No. 72. 9. United Nations VENKAT RAO, M. SwAMINATHAN, AND H. A. B. Research Institute for Social Development, Geneva. PARPIA. 1972. Supplementary value of yeast PANWAR, B. S., B. V. SINGH, AND V. K. GUPTA. 1971. grown on petroleum hydrocarbons to poor diets Soil and foliar applications of urea on "Kalyan based on Kaffir corn and wheat. Plant Foods Hum. Sona" variety of wheat (Triticum aestirum L.). Nutr. 2(3-4): 167-170. Indian J. Agric. Sci. 41(5): 452--455. NATIONAL ACADEMY OF SCIENCES-NATIONAL PARRISH, J. A., AND R. J. BOLT. 1963. Effect of RESEARCH COUNCIL. Food and Nutrition Board. raw wheat germ on nutrition of the chicken. Nature, 1963. Evaluation of Protein Quality. Rep. Int. London 199: 398-399. Conf. Comm. on Protein Malnutrition. Publ. 1109. PARTHASARATHY, H. N., T. R. DORAISWAMY, M. National Research Council, Washington, D.C. 83 p. PANEMANGALORE, M. NARAYANA RAO, B. S. NATIONAL ACADEMY OF SCIENCES-NATIONAL CHANDRASEKHAR, M. SWAMINATHAN, A. SREENI­ RESEARCH COUNCIL. 1964. Recommended Di­ VASAN, AND V. SUBRAHMANYAN. 1963. The effect etary Allowances. Pub. No. 1146. Washington, of fortification of processed soya flour with DI.­ D.C. methionine hydroxy analogue or DL-methionine on NATIONAL RESEARCH COUNCIL. 1969. United the digestibility, biological value, and net protein States-Canadian Tables of Feed Composition. Publ. utilization of the proteins as studied in children. No. 1684. National Research Council, National Can. J. Biochem., 42(3): 377-384. Academy of Sciences, Washington, D.C. PARTRIDGE, J. R. D., AND C. F. SHAYKEWICH. 1972. NELSON, 0. E. 1969. Genetic modifications of pro­ Effects of nitrogen, temperature, and moisture regime tein quality in plants. Adv. Agron. 21: 171-194. on the yield and protein content of Neepawa wheat. NELSON, 0. E., E. T. MERTZ, AND L. S. BATES. 1965. Can. J. Soil Sci. 52(2): 179-185. Second mutant gene affecting the amino acid PEERS, F. G. 1953. The phytase of wheat. Biochem. pattern of maize endosperm proteins. Science 150: J. 53(1): 102-110. 1469-1470. PELLETT, P. L. 1973. Methods of protein evalua­ NUNNIKHOVEN, R., AND E. J. BIGWOOD. 1959. tion with rats, I, pp. 225-244. In J. G. W. Porter and Variation in the composition of amino acids in B. A. Rolls (ed.]. Proteins in human nutrition. wheat flour as a function of the percentage extraction Academic Press, London. from wheat grain (in French). Nutr. Dieta. I : PELLETT, P. L., AND A. R. SAGHIR. 1971. Amino acid 177-191. composition of grain protein from wheat and barley NUTRITION RESEARCH LABORATORY. Hyderabad. treated with 2, 4-D. Weed Res. 11(2-3): 182-189. 196 7-68. Annual Report. PENCE, J. W., R. E. FERREL, P. A. HAMMES, AND W. L. 01so, T. 1971. Clinical studies of amino acid forti­ HALEY. 1965. The composition of commercial fication in Japan. 205-219. In N. S. Scrimshaw, and bulgur. Cereal Sci. Today 10(11): 587-589. REFERENCES 173

PENCE,). W.,C.C. NIMMO,ANDF. N. HEPBURN. 1964. PRIMOST, E., G. RITTMEYER, AND H. H. MAYR. 1967. Proteins. p. 227. In I. Hlynka [ed.]. Wheat Chemistry Experiments to improve the straw strength of and Technology. Monograph Series, Vol. III. cereals. III. Field experiments with chlorcholin­ American Association of Cereal Chemists, St. chloride on winter rye (in German). Bodenkultur. Paul, Minnesota. 18: 41-56. PEPLINSKI, A. J., A. C. STRINGFELLOW, AND E. L. PRINGLE, W. J. S., AND T. MORAN. 1942. Phytic GRIFFIN. l 965. Air classification response of acid and its destruction in baking. J. Soc. Chem. durum and hard red spring wheat flours. Northwest. Ind. 61: 108-110. Miller 272(5): 34, 36--37. PROTEIN ADVISORY GROUP. 1971. Need for nutri­ PEREIRA, A. l 963. Studies on wheat protein varia­ tional and food quality guidelines for plant breeders. tion (in Portuguese). Agronom. Lusit. 25(5): 567- Doc. l.16/5. 19th PAG Meet., Geneva. 582. l 972a. The nature and magnitude of the protein PEREIRA, S. M., A. BEGUM, G. JESUDIAN, AND R. problem. PAG Statement No. 3, Geneva. SUNDERARAJ. l 969. Lysine-supplemented wheat l 972b. PAG guideline for preclinical testing of and growth of preschool children. Am. J. Clin. Nutr. world sources of protein. PAG Guideline No. 6, 22(5): 606--61 l. Geneva. PESSI, Y., M. YLANEN, AND J. SYVALAHTl. 1971. l 973. Upgrading human nutrition through the The effect of fertilization technique on the grain improvement of food legumes. PAG Statement crop of cereals, primarily on the protein content. No. 22, Geneva. Acta Agra!. Fenn. 123: 206--216. PYLER, E. J. 1952. Baking Science and Technology. PHANSALKAR, S. V., M. RAMACHANDRAN, AND V. N. Vol. I and II. Seibel Publishing Co., Chicago. PATW ARDHAN. l 957. Nutritive value of vegetable 803 p. proteins. I. Protein efficiency ratios of cereals and RAND, N. T., AND V. K. COLLINS. l 958. Improving pulses and the supplementary effect of the addition cereals with defatted wheat germ. Food Technol. ofa leafy vegetable. Indian J. Med. Res. 45(4): 611- Champaign. 12(1 l): 585-589. 621. RANHOTRA, G. S. l 972. Hydrolysis during bread­ PLATT, B. S., D.S. MILLER, AND P. R. PAYNE. 1961. making of phytic acid in wheat protein concentrate. In J. F. Brock [ed.]. Recent advances in human J. Food Sci. 37(5): 12-13. nutrition with special reference to clinical medicine. l 973. Factors affecting hydrolysis during bread­ J. & A. Churchill Ltd., London. making of phytic acid in wheat protein concentrate. POLANOWSKI, A. l 967. Trypsin inhibitor from rye Cereal Chem. 50(3): 353-357. seeds. Acta Biochim. Pol. 14(4): 389-395. RANHOTRA, G. S., F. N. HEPBURN, AND W. B. BRADLEY. POMERANZ, Y. l 962. The lysine content of bread l 97 l. Supplemental effect of wheat protein con­ supplemented with soya flour, wheat gluten, dry centrate on the protein quality of white wheat flour. yeast and wheat germ. J. Sci. Food Agric. l 3(2): Cereal Chem. 48(6): 699-706. 78-83. RAo,M.N.,ANDJ.M.McLAUGHLAN. 1967. Lysine l97la. [ed.]. Wheat Chemistry and Technology. and methionine availability in heated casein-glucose American Association of Cereal Chemists. Mono­ mixtures. J. Assoc. Off. Anal. Chem. 50(3): 704-707. graph Series, Vol. III. Revised. American Associa­ RAO, P. B. R., H. W. NORTON, AND B. c. JOHNSON. tion of Cereal Chemists. St. Paul, Minnesota. l 964. The amino acid composition and nutritive l 97 lb. Functional characteristics of triticale, value of proteins. V. Amino acid requirements as a a man-made cereal. Wallerstein Lab. Commun. 34, pattern for protein evaluation. J. N utr. 82( l): 88-92. No. 115: 175-186. RAO, S. C., AND L. I. CROY. l 972. Protease and POMERANZ, Y., B. A. BURKHART, AND L. C. MOON. nitrate reductase seasonal patterns and their rela­ l 970. Triticale in malting and brewing. Proc. Am. tion to grain protein production of "high" vs "low" Soc. Brewing Chem. 40-46. protein wheat varieties. J. Agric. Food Chem. 20(6): POMERANZ, Y., AND K. F. FINNEY. 1973. USDA'ers ll38-ll4l. patent process for high-protein bread. Food Technol. REDDY, K. R., AND R. S. RAO. l 968. Studies on the 27(1): 12-13. effect of soil and foliar application of nitrogen on POMERANZ, Y., AND J. A. SHELLENBERGER. 1971. wheat. Indian J. Sci. Ind. 2(4): 173-176. Bread Science and Technology. Avi Publishing Co., REDDY, V. l 971. Lysine supplementation of wheat Westport, Connecticut. 262 p. and nitrogen retention in children. Am. J. Clin. Nutr. PORTER, J. G. W., AND B. A. ROLLS. 1973. Proteins 24(10): l 246--1249. in human nutrition. Academic Press, London. REINHOLD, J. G. 1971. High phytate content of 560 p. rural Iranian bread: a possible cause of human zinc PRICE, S. A. l 950. The amino acid composition of deficiency. Am. J. Clin. Nutr. 24(10): 1204-1206. whole wheat in relation to its protein content. RILEY, R., AND J. A. D. Ew ART. l 970. The effect of Cereal Chem. 27(1): 73-74. individual rye chromosomes on the amino acid 174 MONOGRAPH IDRC-02le

content of wheat grains. Genet. Res. Camb. 15: SAUNDERS, R. M., AND G. 0. KOHLER. 1972. In 209-219. vitro determination of protein digestibility in wheat ROACH, A. G., P. SANDERSON, AND D. R. WILLIAMS. mill feeds for monogastric animals. Cereal Chem. 1967. Comparison of methods for the determina­ 49(1): 98-103. tion of available lysine value in animal and vegetable SCHIAFFINO, S. S. 1959. Report on microbiological protein sources. J. Sci. Food Agric. 18(7): 274-278. method for amino acids. J. Assoc. Off. Agric. Chem. ROBBINS, G. S., ANDY. POMERANZ. 1971. Proc. Am. 42: 525-527. Soc. Brewing Chemists (in press). SCHLEHUBER,. A. M., AND B. B. TUCKER. 1967. ROBINSON, D. W., AND R. SAGEMAN. 1968. Amino Culture of wheat, pp. 117-179. In K. S. Quisenberry, acid composition of South African and Australian and L. P. Reitz [ed]. Wheat and wheat improvement. wheat varieties as a function of their nitrogen American Society of Agronomy Inc., Madison, content. J. Sci. Food Agric. 19(1): 9-11. Wisconsin. ROGERS, C. G., J. M. MCLAUGHLAN, AND D. G. SCHNEIDER, W., AND H.-J. LANTZSCH. 1971. The CHAPMAN. 1959. Evaluation of protein in foods digestibility of various types of coarse ground rye Ill. A study of bacteriological methods. Can. J. in experiments with sheep and pigs (in German). Biochem. Physiol. 37(11): 1351-1360. Landwirtsch. Forsch. 24(2): 217-221. ROSENBERG, H. R., AND E. L. ROHDENBURG. 1951. SCHRAM, E., S. MOORE, AND E. J. BIGWOOD. 1954. The fortification of bread with lysine. I. The loss of Chromatographic determination of cysteine or lysine during baking. J. Nutr. 45: 593-598. cysteic acid. Biochem. J. 57: 133-137. 1952. 11. The nutritional value of fortified bread. SCHWARZ, R., S. LAUFER, L. LAUFER, AND M. W. Arch. Biochem. Biophys. 37(2): 461-468. BRENNER. 1942. Enrichment of white bread with RozsA, T. A. 1968. Wheat protein concentrate is vitamin B complex. Ind. Eng. Chem. 34: 480-483. here to stay. Bull. Assoc. (US) Operative Millers. Scon, P. M. 1973. Mycotoxins in stored grain, 3054-3059. feeds, and other cereal products. pp. 343-365. In RUCKMAN, J. F., F. P. ZsCHEILE, AND C. 0. QuALSET. R. N. Sinha and W. E. Muir [ed.]. Grain Storage: 1973. Protein, lysine, and grain yields of triticale Part of a System. Avi Publishing Co. Inc., Westport and wheat as influenced by genotype and location. Conn., USA. J. Agric. Food Chem. 21(4): 697-700. SCRIMSHAW, N. S., R. BRESSANI, M. BEHAR, AND F. RUKOSUEV, A. N., AND A. G. SILANT'EvA. 1972. VITERI. 1958. Supplementation of cereal pro­ Amino acid composition of rye grain, sieved rye teins with amino acids II. Effect of amino acid flour, bread flour and bran. Vopr. Pitan. 31(5): supplementation of corn-masa at high levels of 142-145. protein intake on nitrogen retention of young SABISTON, A. R., AND B. M. KENNEDY. 1957. Effect children. J. Nutr. 66: 485. of baking on the nutritive value of proteins in wheat SCRIMSHAW, N. S., AND V. R. YOUNG. 1973. bread with and without supplements of nonfat dry Appraisal of the nutritional significance of new milk and of lysine. Cereal Chem. 34(2): 94-110. proteins for human consumption. Paper to joint SAID, A. K., AND D. M. HEGSTED. 1969. Evalua­ IUPAC/IUFoST Symposium on the contribution tion of dietary protein quality in adult rats. J. Nutr. of chemistry to food supplies. Hamburg. 99: 474-480. SEAMAN, W. L. 1971. Ergot of grains and grasses. SANAHUJA, J. C. 1971. Implications of amino acid Pub!. 1438. Canadian Department of Agriculture, imbalance studies in rats. pp. 179-182. In N. S. Ottawa. Scrimshaw, and A. M. Altschul [ed.]. Amino acid SEARS, E. R. 1968. Relationships of chromosomes fortification of protein foods. MIT Press, Cam bridge, 2A, 2B and 2D with their rye homoeologue. pp. 53- Massachusetts. USBN 0 262 190915. 61. In Proc. 3rd Int. Wheat Genet. Symp. SANTA MARIA, J. 1969. Enrichment of cereal foods SEELEY, R. D., H. F. ZIEGLER, JR., AND R. J. SUMNER. in Chile with fish protein concentrate. pp. 181-184. 1950. The nutritional value of white bread con­ In M. Milner [ed.]. Protein enriched cereal foods for taining nonviable dried yeast. Cereal Chem. 27(1): world needs. American Association of Cereal 50-60. Chemists, St. Paul, Minnesota, 55104. SELL, J. L., G. C. HODGSON, AND L. H. SHEBESKI. SAUER, W. C. 1972. Availability of amino acids 1962. Triticale as a potential component of chick from barley, wheat, triticale and soyabean meal for rations. Can. J. Anim. Sci. 42: 158-166. growing pigs. M.Sc. Thesis. University of Manitoba, SELL, J. L., AND R. L. JoHNSON. 1969. Feeding Winnipeg. value of triticale for turkeys and hens. North SAUNDERS, R. M., M. A. CONNOR, R. H. EDWARDS, Dakota Farm Res. Bimon. Bull. 26(5): 6- 7~ AND G. 0. KoHI.ER. 1972. Enzymatic processing SEN, D. P., T. S. SAYANARAYANA RAO, S. B. KADKOL, of wheat bran: effects on nutrient availability. M. A. KRISHNASWAMY, S. VENKAT RAO, AND N. L. Cereal Chem. 49(4): 436-442. LAHIRY. 1969. Fish protein concentrate from REFERENCES 175

Bombay-Duck (Harpodonn nehereus) fish: Effect of yield of dwarf wheat varieties. Indian J. Agric. Sci. processing variables on the nutritional and organo­ 41(11): 952-958. leptic qualities. Food Technol. 23(5): 683-688. SINGH, S. D., AND J. V. PRASAD. 1966. Effect of SETHI, G. S., AND P. P. SINGH. 1972. Interrelation­ nitrogen and phosphorus fertilization on yield and ship of quantitative traits with grain yield in Triticale. protein content of wheat. Allahabad Farmer 40(1): Indian J. Agric. Sci. 42(4): 281-285. 1-8. SHAMMAS, E., AND W. H. ADOLPH. 1954. Nutritive SISODIA, N. S. 1973. Current status of triticale value of parboiled wheat used in the Near East. breeding in India. Report to CIMMYT-University J. Am. Diet. Assoc. 30~ 982-983. of Manitoba, Triticale Workshop, El Batan, Mexico. SHANDS, H. L. 1969. Rye. pp. 118-149. In S. A. Oct. 1-4, 1973. Matz [ed.]. Cereal Science. Avi Publishing Co. Inc., SMITH, W. H. 1972. Biscuits, crackers and cookies. Westport, Conn., USA. Vol. I. Technology, production and management. SHARMA, R. B., G. P. VERMA, AND S. S. KHANNA. 737 p. Vol. 2. Recipes and formulations. 358 p. 1972. Influence of different moisture regimes and Applied Science Publishers Ltd., London. methods of phosphate application on growth and SNYDERMAN, S. E., P. M. NORTON, D. I. Fowl.ER, AND nutrient absorption by wheat ( Triticwn aestirum L.). L. E. HOLT, JR. 1959. The essential amino acid Indian J. Agric. Sci. 42(1): 52-56. requirements of infants: lysine. Am. J. Diseases SHAW, W. C., C. J. WILLARD, AND R. L. BERNARD. Children 97: 175-185. 1955. Effect of 2,4-dichlorophenoxyacetic acid on SOMIN, V. I. 1970. Amino acids composition of wheat, oats and barley. Res. Bull. Ohio Agric. Exp. different rye varieties prevalent in the USSR (in Stn. 761. Russian). Vopr. Pitan. 29(3): 48-54. SHENK, J. S., F. C. ELLIOTT, AND J. W. THOMAS. 1970. SOSULSKI, F. W., E. A. PAUL, AND W. L. HUTCHEON. Meadow vole nutrition studies with semisynthetic 1963. The influence of soil moisture, nitrogen diets. J. Nutr. 100 (12): 1437-1446. fertilization, and temperature on quality and amino SHEPHERD, A. D., R. E. FERREL, N. BELLARD, AND acid composition of Thatcher wheat. Can. J. Soil J. W. PENCE. 1965. Nutrient composition of Sci. 43: 219-228. bulgur and lye-peeled bulgur. Cereal Sci. Today. SPIES, J. R. 1967. Determination of tryphophan in I 0(11): 590-592. protein. Anal. Chem. 39: 1412-1416. SHERROD, L. B. 1972. Nutritive value of selected SPIES, J. R., AND D. c. CHAMBERS. 1949. Chemical triticales and wheats. J. Anim. Sci. 34(5): 909. determination of tryptophan in proteins. Anal. SHIMADA, A. S., AND T. R. CLINE. 1972. Nutritional Chem. 21 : 1249-1266. evaluation of triticale. J. Anim. Sci. 35(5): 1110. SRIVASTAVA, K. N., V. PRAKASH, R. DE, AND M. S. SHIMADA, A. S., R. L. MARTINEZ, AND F. 0. BRA VO. NAIK. 1971. Improvement of the quality of pro­ 1971. Studies on the nutritive value of triticale for teins of ""S-227" wheat by nitrogen fertilization growing swine. J. Anim. Sci. 33(6): 1266-1269. under rain-fed conditions. Indian J. Agric. Sci. SHORROCK, C., AND J.E. FORD. 1973. An improved 41(3): 202-205. procedure for the determination of available lysine STEELE, B. F., H. E. SAUBERLICH, M. S. REYNOLDS, AND and methionine with Tetrahymena. pp. 207-223. C. A. BAUMANN. 1949. Media for Leuconostoc In J. W. G. Porter and B. A. Rolls [ed.]. Proteins in mesenteroides P-60 and Leuconostoc citrororum human nutrition, Academic Press, London. 8081. J. Biol. Chem. 177: 533-544. STEIN, W. H., AND S. MOORE. 1951. Electrolytic SHOUP, F. K., Y. POMERANZ, AND C. W. DEYOE. 1966. desalting of amino acids. Conversion of arginine to Amino acid composition of wheat varieties and ornithine. J. Biol. Chem. 190: 103-106. flours varying widely in bread-making potentialities. STILLINGS, B. R., V. D. SIDWELL, AND 0. A. HAMMERLE. J. Food Sci. 31(1): 9~101. 1971. Nutritive quality of wheat flour and bread SHYAMALA, G., AND B. M. KENNEDY. 1962. Protein supplemented with either fish protein concentrate or value of chapatis and puris. J. Am. Diet. Assoc. lysine. Cereal Chem. 48(3): 292-302. 41: 115-118. STOTHERS, S. C., AND L. H. SHEBESKI. 1965. The SIDwELL, V. D. 1967. U.S.Q.M. Associates Activi­ nutritive value of triticale for growing swine. ties Report 19: 118-124. pp. 17-18. In 14th Rep. on Livestock Research. SIHLBOM, E. 1962. Amino acid composition of Dept. Animal Sci. University of Manitoba, Canada. Swedish wheat protein. Acta. Agric. Scand. 12: STOTT, J. A., AND H. SMITH. 1966. Microbiological 148-156. assay of protein quality with Tetrahymena pyriformis SIMMONDS, D. H. 1962. Variations in the amino acid W. 4. Measurement of available lysine, methionine, composition of Australian wheats and flours. Cereal arginine and histidine. Br. J. Nutr. 20: 663-673. Chem. 39(6): 445-455. STRINGFELLOW, A. C., AND A. J. PEPLINSKI. 1966. SINGH, N. P., AND N. G. DASTANE. 1971. Effects of Air classification of Kansas hard red winter wheat moisture regimes and nitrogen levels on growth and flours. Northwest. Miller. 370(6): 19-20, 22. 176 MONOGRAPH IDRC-02le

STRINGHAM, E. W. 1968. Triticale in the finishing whole meal chapatis and leavened bread. J. Agric. of beef cattle. Talk presented at Beef Cattle Field Food Chem. 20(1): 116-118. Day, Brandon, Manitoba, Canada. TARA, K. A., M. S. UsHA, C. PRABHAVATI, AND G. S. STROMNAES, A. S., AND B. M. KENNEDY. 1957. BAINS. 1971. Vitamin, mineral, colour grade and Effect of baking on the nutritive value of proteins in limiting amino acid contents of Indian commercial rye bread with and without supplements of non fat flours. Res. Indus. 16(3): 196-202. dry milk and of lysine. Cereal Chem. 34(3): 196--200. TASKER, P. K., T. R. DORAISWAMY, M. NARAYANARAO, STUBER, C. W., V. A. JOHNSON, AND J. W. SCHMIDT. M. SwAMINATHAN, A. SREENIVASAN, AND V. 1962. Grain protein content and its relationship SUBRAHMANYAN. 1962. The metabolism of nitro­ to other plant and seed characters in the parents and gen, calcium and phosphorus in undernourished progeny of a cross of Triticum aestimm L. Crop children. 8. The metabolism of nitrogen, calcium Science, 2(6): 506--508. and phosphorus, and the digestibility coefficient SULLIVAN, B. l 967a. Wheat based products for and biological value of the proteins and the net pro­ world use. Cereal Sci. Today 12(10): 446--448, 462. tein utilization on poor Indian diets based on rice, l 967b. Wheat-based products for world use in maize or a mixture of rice and maize. Br. J. Nutr. United States aid programs. Am. Miller Processor 16: 361-368. 95(6): 11-12, 30. TAYLOR, A. C., AND A. R. GILMOUR. 1971. Wheat SULTAN, W. J. 1969. Practical baking. 2nd ed. Avi protein prediction from climatic factors in southern Publishing Co., Westport, Connecticut. 492 p. New South Wales. Aust. J. Exp. Agric. Anim. SURE, B. 1948. The nature of the supplementary Hush. 11 : 546--549. value of the proteins in milled corn meal and TAYLOR, Y., V. R. YOUNG, AND N. S. SCRIMSHAW. milled wheat flour with dried food yeasts. J. Nutr. 1972. Lysine supplementation of wheat gluten at 36: 59-63. restricted caloric level in young men. Fed. Proc. 1954. Relative nutritive values of proteins in 31(2): 717. whole wheat and whole rye and effect of amino acid TEERI, A. E., W. VIRCHOW, AND M. E. LOUGHLIN. supplements. J. Agric. Food Chem. 2(22): 1108- 1956. A microbiological method for the nutri­ 1110. tional evaluation of proteins. J. Nutr. 59(4): 587- SwAMINATHAN, M. S., A. AusnN, A. K. KAUL, AND 593. M. S. NAIK. 1969. Genetic and agronomic enrich­ TENNENHOUSE, A. N., AND L. J. LACROIX. 1972. ment of the quantity and quality of proteins in cereals Effects of (2-chloroethyl) trimethylammonium chlo­ and pulses. pp. 71-86. In Proceedings of a panel ride (CCC) on certain agronomic traits of oats and meeting on new approaches to breeding for plant triticale. Can. J. Plant Sci. 52: 559-567. protein improvement. International Atomic Energy THOMAS, K. 1909. Arch. Anal. Physiol. Abstr. 219. Agency, Vienna. THOMPSON, B. N. 1971. An investigation of the SZABO, L. GY. 1972. The germination physiology effects of chlorine treatment on the cake baking of triticale. Acta Agron. Acad. Sci. Hung. 21(1-2): properties of triticale flour. M.Sc. Thesis. University 219-222. of Manitoba, Canada. SZKILLADZIOWA, W. l 960a. Biological value of pro­ THOMPSON, B. N., AND F. M. VAISEY. 1971. An tiens in rye and wheat flour of different extraction investigation of the effects of chlorine treatment on rates, estimated from growth trials (in Polish). Rocz. the cake baking properties of Triticale flour. Cereal Panstw. Zakt. Hig. 11: 191-198. Sci. Today 16(9): 304 (Abstr.). l 960b. The biological value of protein in rye and wheat flour of various extraction rates estimated TKACHUK, R. 1969. Nitrogen-to-protein conversion by means of Bender and Miller's method (in Polish). factors for cereals and oilseed meals. Cereal Chem. Rocz. Panstw. Zakt. Hig. 11 (4): 295--301. 46(4): 419-423. 1961. Nutritive value of rye flour proteins esti­ TKACHUK, R., AND G. N. IRVINE. 1969. Amino acid mated by a growth method of rats given a low level of compositions of cereals and oilseed meals. Cereal protein (in Polish). Rocz. Panstw. Zakt. Hig. 12(6): Chem. 46: 206-218. 563-568. TOEPFER, E.W., M. E. HEWSTON, F. N. HEPBURN, AND 1962. Protein supplementation of rye flours J. H. TuLLOss. 1969. Nutrient composition of with some other food products (in Polish). Rocz. selected wheats and wheat products. I. Description Panstw. Zakt. Hig. 13: 415-425. of samples. Cereal Chem. 46(5): 560-567. SZMELCMAN, S., AND K. GUGGENHEIM. 1967. TOEPFER, E. W., M. M. POLANSKY, J. F. EHEART, Availability of amino acids in processed plant H. T. SLOVER, E. R. MORRIS, F. N. HEPBURN, AND protein foodstuffs. J. Sci. Food Agric. 18: 347-350. F. W. QUACKENBUSH. 1972. Nutrient composi­ TARA, K. A., M. S. M. UsHA, AND G. S. BAINS. 1972. tion of selected wheats and wheat products. XI. Effects of lysine on dough and protein quality of Summary. Cereal Chem. 49(2): 173-186. REFERENCES 177

TRZEBSKA-JESKE, I., AND W. MORKOWSKA-GLUZINSKA. vitamins, and gross energy. Cereal Chem. 44(1): 1963. Contents of nitrogen and some essential 48-60. amino acids in Polish rye grain harvested in 1959 (in WALL, J. S., C. JAMES, AND J. F. CAVINS. 1971. Polish). Rocz. Panstw. Zakt. Hig. 14(2): 153-166. Nutritive value of protein in hominy feed fractions. TSEN, C. C., W. J. HOOVER, AND E. P. FARRELL. 1973. Cereal Chem. 48(4): 456-465. Baking quality oftriticale flours. Cereal Chem. 50( 1): wANG, c. C., AND D. R. GRANT. 1969. The pro­ 16--26. teolytic enzymes in wheat flour. Cereal Chem. 46(5): TsUNENAKI, K. 1968. Origin and phylogenetic dif­ 537-544. ferentiation of common wheat revealed by com­ WANG, H. L., D. I. RUTTLE, AND C. W. HESSELTINE. parative gene analysis. pp. 71-85. In Proc. 3rd Int. 1968. Protein quality of wheat and soybeans after Wheat Genet. Symp. Rhizopus oligosporus fermentation. J. Nutr. 96(1): UNITED NATIONS. 1968. International action to avert 109-114. the impending protein crisis. Report to the Economic WEBER, C. W., J. 0. NORDSTROM, AND B. L. REID. and Social Council of the United Nations. New 1972. Grain sorghum, wheat and triticale in laying York. hen diets. Poult. Sci. 51(5): 1885. 1971. Strategy statement on action to avert the WEBER, c. W., AND B. L. REID. 1971. Utilization of protein crisis in the developing countries. Report of triticale and various wheat varieties by laying hens. the panel of experts on the protein problem con­ Poul. Sci. 50(5): 1644. fronting developing countries. UN Headquarters, 1972. Protein evaluation of Mexican wheat and New York. triticale varieties by the use of young mice. Fed. UNRAU, A. M., AND B. S. JENKINS. 1964. Investiga­ Proc. Fed. Amer. Soc. Exp. Biol. 31 (2): A690. tions on synthetic cereal species. Milling, baking and WENCKERT, E., E.-M. LOESER, S. N. MAHAPATRA, F. some compositional characteristics of some triticale SCHENKER, AND E. M. WILSON. 1964. Wheat and parental species. Cereal Chem. 41 (5): 365-375. bran phenols. J. Org. Chem. 29: 435-440. UPRETI, D. C., ANDY. P. ABROL. 1971. Studies on WESTERMAN, B. D., B. OLIVER, AND E. MAY. 1954. chemical constituents of triple gene dwarf wheat Improving the nutritive value of flour. VI. A com­ strains. Bull. Grain Technol. 9( 1): 20-25. parison of the use of soya flour and wheat germ. URIE, A., AND J. H. HULSE. 1952. The science, raw J. Nutr. 54(2): 225-235. materials and hygiene of baking. MacDonald and WESTERMAN, B. D., F. ROACH, AND M. STONE. 1952. Evans Ltd., London. 458 p. Improving the nutritive value of flour. V. The effect VAISEY, M., AND A. M. UNRAU. 1964. Chemical of the use of defatted wheat germ. J. Nutr. 47(1): constituents of flour from cytologically synthesized 147-158. and natural cereal species. J. Agric. Food Chem. WIDDOWSON, E. M., AND R. A. MCCANCE. 1954. 12( 1): 84-86. Studies of the nutritional value of bread and the VALLEJO, J.M., S. DE LA PLAZA, M. SALTO, AND F. G. effect of variation in the extraction rate of flour on OLMEDO. 1969. Utilization of hexaploid triticale. the growth of undernourished children. Med. Res. II. Rheological and baking tests (in Spanish). An. Counc. Spec. Rep. No. 287. H.M. Stationery Office, Invest. Agron. 18(4): 371-381. London. VILLEGAS, E., C. E. McDONALD, AND K. A. GILLES. WIERINGA, G. W. 1967. On the occurrence of 1970. Variability in the lysine content of wheat, growth inhibiting substances in rye. Pub!. 156. rye and triticale proteins. Cereal Chem. 47(6): Institute for Research on Storage and Processing 746--757. of Agricultural Produce. Wageningen, The Nether­ VLASYUK, P. A., AND L. N. KURINNAYA. 1971. lands. 68 p. Change of the fractional composition of proteins of 1967 /68. Considerations on the use of rye winter wheat grain under application of fertilizer in and wheat as feed (in Dutch). pp. 157-160. In a crop rotation system. Agrokhim. 9: 20-23. Year book 1967 /68. Institute for Research on VOGEL, H.J., AND R.H. VOGEL. 1967. Regulation Storage and Processing of Agricultural Produce. of protein synthesis. Ann. Rev. Biochem. 36: 519- Wageningen, The Netherlands. 538. WILLIAMS, P. C. 1966. Reasons underlying varia­ VON LOESECKE, H. W. 1946. Controversial aspects: tions in the protein content of Australian wheat. Yeast in human nutrition. J. Am. Diet. Assoc. 22: Cereal Sci. Today 11(8): 332-335, 338. 485-493. WINICK, M., AND P. Rosso. 1969. Head circum­ WAGGLE, D. H., M.A. LAMBERT, G.D. MILLER, E. P. ference and cellular growth of the brain in normal FARRELL, AND c. W. DEYOE. 1967. Extensive and marasmic children. J. Pediatr. 74: 774-778. analyses of flours and millfeeds made from nine 1970. Malnutrition and cellular growth in the different wheat mixes. II. Amino acids, minerals, brain. pp. 531-538. In Proc. 8th Inter. Cong. on 178 MONOGRAPH IDRC-02le

Nutrition, Prague 1969. Excerpta Medica Amster­ some emulsifiers and stabilizers and certain other dam. ISBN 90 219 0139 0. substances. WHO Technical Report "Series 373. WINSTON, J. J. 1971. Macaroni, Noodles and Pasta FAO Nutrition Meetings Ser. 43. Tenth report of Products. I. N. Publishing Corp., 233 Lexington the Joint FAO/WHO Expert Committee on Food Avenue, New York 10016. Additives. 47 p. WOMACK, M., M. W. MARSHALL, AND J. S. SUMMERS. 1970. Specifications for the identity and purity 1954. Nutritive value of bread and cookies con­ of food additives and their toxicological evaluation: taining cottonseed flour. J. Agric. Food Chem. some food colours, emulsifiers, stabilizers, anti­ 2(3): 138-140. caking agents, and certain other substances. WHO WORLD HEALTH ORGANIZATION. 1957. General Technical Report Series 445. FAO Nutrition Meet­ principles governing the use of food additives. WHO ings Series 46. Thirteenth report of the Joint FAO/ Technical Report Series 129. FAO Nutr. Meetings WHO Expert Committee on Food Additives. 31 p. Rep. Series 15. First report of the Joint FAO/WHO 1972. Evaluation of Food Additives. Some en­ Expert Committee on Food Additives. 22 p. zymes, modified starches, and certain other sub­ 1958. Procedures for the testing of international stances: toxicological evaluation and specifications food additives to establish their safety in use. WHO and a review of the technological efficiency of some Technical Rep. Series 144. FAO Nutr. Meetings antioxidants. WHO Tech. Rep. Ser. 488. FAO Rep. Series 17. Second report of the Joint FAO/ Nutrition Meetings Series 50. Fifteenth rep. of the WHO Expert Committee on Food Additives. 19 p. Joint FAP/WHO Expert Committee on Food 1961. Evaluation of the carcinogenic hazards of Additives. 41 p. food additives. WHO Technical Rep. Series 220. 1973. Energy and protein requirements. WHO FAO Nutr. Meetings Rep. Series 29. Fifth Rep. of Tech. Rep. Series 522. FAO Nutrition Meetings the Joint FAO/WHO Expert Committee on Food Rep. Series 52. Report of a Joint FAO/WHO Ad Additives. 33 p. Hoc Expert Committee. 1962. Evaluation of the toxicity of a number of WOSTMANN, B. 1950. The cystine content of wheat antimicrobials and antioxidants. WHO Technical flour in relation to dough properties. Cereal Chem. Rep. Series 228. FA 0 N utr. Meetings Rep. Series 31. 27: 391-397. Sixth Rep. of the Joint FAO/WHO Expert Com­ WRICKE, G. 1962. Z. Pflanzenzuecht. 47: 92-96. mittee on Food Additives. 104 p. YAMASAKI, K. 1968. L-lysine enrichment of bread. 1964. Specifications for the identity and purity Food Ind. Asia 1(4): 14-16. of food additives and their toxicological evaluation: YANEZ, E., D. BALLESTER, I. BARJA, N. PAK, A. REID, emulsifiers, stabilizers, bleaching and maturing E. TRABUCCO, I. PENNACCHIOTTI, L. MASSON, M. A. agents. WHO Technical Report Series 281. FAO MELLA, J. VINAGRE, 0. CERDA, H. SCHMIDT­ Nutr. Meetings Rep. Series 35. Seventh Rep. of the HEBBEL, J. V. SANTA MARIA, AND G. DONOSO. Joint FAO/WHO Expert Committee on Food 1967. Biological study of new protein sources for Additives. 189 p. human consumption (in Spanish). Nutr. Bromatol. 1965a. Protein requirements. Joint FAO/WHO Toxicol (6): 85-98. Expert Group. WHO Tech. Rep. Ser. No. 301. YANEZ, E., AND J. M. MCLAUGHLAN. 1970. The FAO Nutr. Meetings Rept. Series 37. effect of level of protein on protein quality of l 965b. Specifications for the identity and purity lysine-deficient foods. Can. J. Physiol. Pharmacol. of food additives and their toxicological evaluation: 48(3): 188-192. food colours and some antimicrobials and anti­ YANEZ, E., H. WULF, 0. BALLESTER, N. FERNANDEZ, oxidants. WHO Technical Rep. Ser. 309. FAO Nutr. V. GATTAS, AND F. MONCKEBERG. 1973. Nutritive Meetings Rep. Ser. Eighth rep. of the Joint FAO/ value and baking properties of bread supplemented WHO Expert Committee on Food Additives. 25 p. with Candida uti/is. J. Sci. Food Agric. 24: 519-525. 1966. Specifications for the identity and purity YANG, S. P., AND F. F. AL-NOURI. 1962. Nutritive of food additives and their toxicological evaluation: value of the protein of parboiled wheat. Fed. Proc. some antimicrobials, antioxidants, emulsifiers, stabi­ 21: 395. lizers, flour-treatment agents, acids and bases. WHO YANG, S. P., H. E. CLARK, AND G. E. VAIL. 1961. Technical Rep. Ser. 339. FAO Nutrition Meetings Effects of varied levels of a single daily supplement Ser. 40. Ninth rep. of the Joint FAO/WHO Expert of lysine on the nutritional improvement of wheat Committee on Food Additives. 24 p. flour proteins. J. Nutr. 75(2): 241-246. 1967a. Procedures for investigating intentional YONG, F. C., AND A. M. UNRAU. 1964. Influence of and unintentional food additives. WHO Tech. Rep. alien genome combinations on protein synthesis in Ser. 348. Rep. of a WHO Scientific Group. 25 p. cereals. Can. J. Biochem. 42: 1647-1657. l 967b. Specifications for the identity and purity YOUNG, V. R.,AND N. S. SCRIMSHAW. 1971. Clinical of food additives and their toxicological evaluation: studies in the United States on the amino acid REFERENCES 179

fortification of protein foods. pp. 248-268. In N. S. ZIEGLER, E., AND E. N. GREER. 1971. Principles of Scrimshaw and A. M. Altschul [ed.]. Amino acid milling, pp. 115-199. In Y. Pomeranz [ed.]. Wheat fortification of protein foods. MIT Press, Cam­ Chemistry and Technology. Monograph Series. Vol. bridge, Massachusetts. 111 revised. American Association of Cereal Chem­ 1972. The nutritional significance of plasma and ists, St. Paul, Minnesota. urinary amino acids, pp. 541-568. In E. J. Bigwood Zn.LINSKY, F. J. 1973. Triticale breeding and re­ [ed.]. Protein and amino acid functions. International search at CIMMYT. Research Bulletin No. 24. Centro Encyclopaedia of Food and Nutrition. Vol. 11. Internacional de Mejoramiento de Maiz y Trigo, Pergamon Press, Oxford. Mexico 6. Zn.LINSKY. F. J., AND N. E. BoRLAUG. 197la. Pro­ ZHEBRAK, E. A., AND N. M. Ko1.CHIN. 1968. Pro­ gress in developing triticale as an economic crop. tein composition of diploid and tetraploid rye (in Res. Bull. No. 17. Centro Internacional de Mejor­ Russian). Sel'sk. Biol. 3: 439-441. amiento de Maiz y Trigo, Mexico 6. 27 p. ZENTNER, H. 1961. Influence of glycine and lysine 1971 b. Search for a new food source for man: on dough fermentation. J. Sci. Food Agric. 12(12): triticale research in Mexico. Agric. Sci. Review 812-817. 9(4): 28-35. Index

Additives 103 phytic acid 97 Aftatoxins I 02 processing 21, 109, 124 Agronomic effects 16, 18, 44, 69, 81, 89, 92 protein 9, 27, 107 Air classification 112, 118 supplementation 22, I 09, 134 Alimentary pastes 125, 130, 137, 146 toxic factors I 0 I Amino acids Cereal products analytical methods 29, 37 protein supplement 22, 143 availability 30, 120 Chapatis 129, 139, 141, 142 baking effect 127 Chemical score 29 imbalance 23 Chorleywood Bread Process 128 "non-essential" 142 CIMMYT 7 recommended levels 12 Clariceps purpurea (see Ergot) reference patterns 30, 37 Conversion factors 28 rye 12, 45, 88, 111, 124 Cottonseed protein 149 synthetic supplements 22, 134 triticale 12, 17, 45, 111, 124 Dye-binding methods 35 wheat 12,45, 71, 73, 78, 82, 88, Ill wheat flour 12, 113, 118 Analytical methods 28, 37 Egg Animal feed protein supplement 145 ergot IOI Environmental effects 18, 44, 48, 69, 74, 89 resorcinols 95 Enzyme inhibitors 99 triticale 17, 53 Ergot 20, 62, 65, 67, IOI, 103 Evaluation methods 12, 27 Baking 109, 127, 134, 136, 139, 141, 143 Baking powder biscuits 130 Fertilizers effect 69, 81 Barley 48, 57, 64, 67, 69 Fish protein 23, 139, 152 Bibliographies 41 Biological assays methodology 32, 36, 38 Genetic factors 16, 18, 41, 70 Bran Germ protein supplement 143 protein supplement 143 Breakfast cereals 125, 131 Gluten Brewing 125 protein supplement 143 Bulgur 131 Groundnut meal protein supplement 139 Growth-depressing factors I 00 Cake 124, 130 Growth slope assay 34, 37, 38 Canadian International Development Agency 8 Cattle feeding 67, 102 CCC 69, 93 Human feeding trials 54, 117, 119, 128, 135, 138, 144, Cereals (see also Rye, Triticale, Wheat) 148, 150, 156 calorie source 9 growth-depressing factors I 00 milling 105 International Development Research Centre 8 mycotoxins 102 International Union of Food Science and nitrogen conversion factors 28 Technology 21 nitrogen determination 28, 37 Isoleucine nutritional inhibitors 95, I 03 protein supplement 141

181 182 MONOGRAPH IDRC-02le

Legumes 9, 23, 28, 149 Rat assays Lysine (see also Amino acids) methodology 32, 38 baking loss 127, 134, 136, 139, 141 rye 54, 88, 90, 95, 100, 122, 128, 129 genetic control 42, 71 triticale 54 protein supplement 134, 142, 145, 150, 154 wheat 54, 77, 86, 95, JOO wheat flour 116, 120, 127, 129, 132, 134, 136, 142, Maize 42, 54, 60, 66 145, 149, 151, 156 Malting 125 Reference patterns 30, 37 Manitoba, University of 7 Resorcinols 19, 96 Meadow vole (see Microtus pennsylvanicus) Rhizopus oligosporus fermentation 133 Methionine Roasting 127 Rye (see also Cereals) protein supplement 134, 137, 141, 150 Microorganisms aftatoxins 102 agronomy 89, 92 protein supplement 23, 151 Microtus pennsylvanicus assays 36, 56 amino acids 12, 45, 88, 111, 124 Microwave baking 129 biological quality 54, 59, 88, 90, 92, 93, 95, JOO, Milk 102 protein supplement 23, 134, 136, 139, 145 environmental effects 48 Milling 20, I 05, 110 enzyme inhibitors 99 "Mote" 127 ergot 20, I 0 I Mouse assays 36, 56, 96, 117, 129 genetic factors 16, 41 Mycotoxins 20, 102 growth-depressing factors I 00 milling 21,110,IIl,121 nutritional inhibitors 19, 95 Net protein ratio phytic acid 19, 97 methodology 33, 37, 38 processing 21, 128 Net protein utilization production data 8 methodology 33 protein 11, 45, 88, 90 Nitrogen proteolytic activity 111 conversion factors 28 resorcinols 19, 95 determination 28, 37 reviews 41 plant metabolism 70 trypsin inhibitors 20, 99 Nutritional inhibitors 19, 95 varietal effects 16, 19, 41, 88, 90 varieties 45, 48, 51, 56, 88, 90, 95, 111 Rye flour Phytic acid 19, 97 air classification 112 Poultry feeding 59, 93, I 00, 132, 148 amino acids 124 Processing 21, 124 baking 128 Protein biological value 122, 128 analysis 28, 37 processing 128 biological evaluation 32, 36, 38 supplementation 134, 145 cereal IO, 18, 45, 70, 72, 77, 81, 88, 90 chemical score 30 classification 43 Scones 130 dietary importance IO Sheep feeding 67, 102 evaluation methods 12, 27 Skim milk powder (see Milk) seed distribution I 05 Slope ratio assay 33 sources 9 Sorghum 63, 65, 68 supplementation 22, 109, 134 Soya 58, 61, 64, 139, 147, 150 Protein efficiency ratio Steaming 127, 133 methodology 32, 34, 38 Supplementation Proteolytic activity I I I amino acids 22, 134 Pulses benefits and costs 23 protein supplement 149 cereal by-products 22, 143 cereal protein 22, I 09, 134 cottonseed protein 149 Rapid screening methods 34 fish protein 22, 139, 152 HULSE/LAING: PROTEINS OF TRITICALE 183

legumes 22, 149 Tryptophan microorganisms 22, 151 protein supplement 141 milk 22, 134, 136, 139, 145 Trypsin inhibitors 20, 99 soya 147, 150 vegetables 149 yeast 151 Valine Swine feeding 64, 87, 92, 95, 102 protein supplement 141 Varietal effects cereal protein 16, 47, 72, 88, 90 Tempeh 133 Vegetables Threonine protein supplement 149 protein supplement 134, 136, 137, 141, 150, 154 Vole (see Microtus pennsylvanicus) Tolerance levels 103 Triticale (see also Cereals) agronomy 69 Wet-milling 112 amino acids 12, 17, 18,45,46, Ill, 124 Wheat (see also Cereals) biological quality 17, 53 aflatoxins I 02 breakfast cereals 125 agronomy 81, 89 brewing 125 amino acids 12, 45, 71, 73, 78, 82, 88, 111 cattle feeding 67, I 02 biological quality 53, 66, 69, 77, 86, 95, 100 CCC treatment 69 environmental influences 18, 44, 48, 74, 89 chymotrypsin inhibitor 99 enzyme inhibitors 99 CIMMYT-Manitoba project 7, 9 ergot 20, 101 development 7 genetic factors 16, 41, 70 environmental factors 18, 44, 48, 69 grain structure I 05 ergot 20, 62, 65, 67, IOI growth-depressing factors I 00 genetic factors 16, 41 milling 21, 113, 124 human feeding trials 53 nitrogen metabolism 70 malting 125 nutritional inhibitors 19, 95 Microtus pennsylvanicus assays 56 phytic acid 19, 97 milling 20, I 09, 124 processing 21, 127 mouse assays 56, 96 production data 8 mycotoxins 20 protein content 11, 18, 45, 70, 72, 78, 81 nutritional inhibitors 19, 95 proteolytic activity 111 poultry feeding 58, I 01 resorcinols 19, 95 processing 21, 124 reviews 41 production data 8 trypsin inhibitors 20, 99 protein content 11, 45, 69 varietal effects 16, 18, 41, 70, 88 proteolytic activity 111 varieties 45, 48, 51, 56, 61, 63, 70, 72, 74, 83, 87, rat assays 54 88, 101, 102, Ill, 124 research 7 Wheat flour resorcinols 19, 96 air classification 112, 118 reviews 41 amino acids 113, 118 sheep feeding 67, 102 baking 127 solubility characteristics 111 biological quality 116, 120, 127, 131, 134, 144, 151 swine feeding 64 156 trypsin inhibitor 99 Chorleywood Bread Process 128 varietal factors 16, 41 "mote" 127 varieties 45, 48, 52, 61, 110, 125 processing 127 wet-milling 112 supplementation 134 Triticale flour Wheat protein concentrate 98, 144 air classification 112 World Wheat Collection 72, 77, 80 alimentary pastes 128 breadmaking 127 cake making 128 Yeast prepared mixes 127 protein supplement 151